Potential genetic markers for wool quality traits reported by various researchers.
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Hickey is currently Head of the Department of Geography and also Head of the School of the Human Environment at the University College Cork, in addition, he is a Fellow of the Royal Meteorology Society and the Royal Geographical Society.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"17924",title:"Dr.",name:"Kieran",middleName:"Richard",surname:"Hickey",slug:"kieran-hickey",fullName:"Kieran Hickey",profilePictureURL:"https://mts.intechopen.com/storage/users/17924/images/system/17924.jpg",biography:"Dr. Kieran R. Hickey is a Senior Lecturer in Physical Geography in the School of the Human Environment in University College Cork, Rep. of Ireland where he is currently Head of the Department of Geography and also Head of the School of the Human Environment. 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Venkateswarlu",coverURL:"https://cdn.intechopen.com/books/images_new/371.jpg",editedByType:"Edited by",editors:[{id:"58592",title:"Dr.",name:"Arun",surname:"Shanker",slug:"arun-shanker",fullName:"Arun Shanker"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"878",title:"Phytochemicals",subtitle:"A Global Perspective of Their Role in Nutrition and Health",isOpenForSubmission:!1,hash:"ec77671f63975ef2d16192897deb6835",slug:"phytochemicals-a-global-perspective-of-their-role-in-nutrition-and-health",bookSignature:"Venketeshwer Rao",coverURL:"https://cdn.intechopen.com/books/images_new/878.jpg",editedByType:"Edited by",editors:[{id:"82663",title:"Dr.",name:"Venketeshwer",surname:"Rao",slug:"venketeshwer-rao",fullName:"Venketeshwer Rao"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"40793",title:"Identification of Polymorphism in the Keratin Genes (KAP3.2, KAP6.1, KAP7, KAP8) and Microsatellite BfMS in Merino Sheep Using Polymerase Chain Reaction-Single Strand Conformational Polymorphism (PCR-SSCP) Analysi",doi:"10.5772/45762",slug:"identification-of-polymorphism-in-the-keratin-genes-kap3-2-kap6-1-kap7-kap8-and-microsatellite-bfms-",body:'Wool production is a major agricultural industry world-wide, the most important wool-growing countries include Australia, China, New Zealand, South Africa and countries within South America. In Australia for example, the world\'s largest producer of wool accounting for ~ 30% of the world production, wool industry is among the top industries in export revenue. While Australia has long been associated with the production of high-quality wool, the importance of this industry and the value of wool exports have been steadily declining.
The wool industry is faced with many challenges that require innovative solutions. The major competitors to the wool industry, cotton and synthetics, have developed new fibres that meet consumer needs such as being lightweight, soft and easy to care. These competitors have also made better productivity gains than wool, which has resulted in lower prices for all textile products. Today, there is much instability in wool prices, with a major problem facing the industry in faulty wool production. It has been observed that considerable variation exists both within and between fleeces across sheep breeds, as well as within inbred lines of sheep. Since the efficiency of wool processing is dependent on the consistency of wool fibre, it is of prime importance to wool producers that this variation is controlled. The wool characteristics that are of economic importance include fibre diameter (or fineness), grease and clean fleece weight, fleece strength and length, colour, yield, crimp and bulk. For Merino and halfbred wools, fibre diameter is the major factor that contributes to price variation as it significantly influences both fibre processing properties and ultimate product quality. The colour of wool is also important because superior colour (bright and white) can be dyed to the maximum range of shades and consequently is worth more than poorer coloured wool. Furthermore, the quantity of wool is important in overall wool production and in the efficiency of the production system.
For many years, farmers have been using classical selective breeding, where by selection of breeding animals was traditionally based on the phenotype (that is appearance) of the individual animal, a rather slow method of selection. Each animal is assigned a breeding value (BV), which describes the future genetic potential of an animal. The BV is calculated by adjusting phenotype to exclude factors such as birth rank, lambing status and sex in order to give an estimate of the genetic merit. The desired goal of this strategy is the accumulation of “good” forms of genes for that particular trait in the population, over time. This has resulted in many breeds that are commercially important today. The domestic sheep Ovis aries today comprises over 500 different domestic breeds. However, wool characteristics, like many production traits (such as milk yield, growth rate, meat tenderness), do not exhibit simple Mendelian inheritance patterns (recessive or dominant). Instead, they are controlled by not only many genes, but also the interaction of these genes, each having small additive effects on the phenotype observed. Environmental and management factors also play a role. Thus, wool traits are quantitative and show continuous variation in phenotype, a fact that makes it difficult to deduce the genotype of an animal from its phenotype, and to relate genetic variation to differences in the phenotype. In other words, genetic improvement breeding programme select for “phenotypic superior” animals, without the knowledge of the actual genes that are being selected – which I will term as “blind selection” in this paper. Furthermore, other strategies to control environmental factors such as nutrition, time of shearing or mineral supplementation tend to be costly. In addition, wool production traits tend to only be fully expressed when an animal is mature, at least three years old, and therefore genetic progress using phenotypic selection and pedigree information is relatively slow.
The answer to sidestepping this “blind selection”, inaccuracy in describing the genetic potential of an animal and slow progress may lie in identifying specific genetic markers that are associated with wool production traits. Some sheep consistently produce quality or faulty wool, suggesting that genetic factors are an important key in determining wool characteristics. In addition, estimates for the heritability (h2) of most wool traits are generally high (h2 = 0.3 - 0.6), indicating that wool traits are under genetic control and that they can be selected for. A gene is a segment of DNA that provides the genetic information necessary to produce a protein. For almost all of the genes, there are two copies (alleles), one inherited from the mother and the other from the father. In any population of animals, there can be many different alleles. This is termed polymorphism or genetic variation. Polymorphism results from DNA mutation. It is this polymorphism that is taken advantage of, in order to identify genetic markers. A genetic marker for a particular characteristic can be defined as a piece of DNA that directly affects a phenotype and shows polymorphism. It can also be a piece of DNA that is closely linked to another piece of DNA that affects a phenotype. Genetic markers can either be genes or non-functional DNA segments such as microsatellites or minisatellites.
A number of different types of genetic markers are commonly used, including restriction fragment length polymorphisms (RFLPs), microsatellite and minisatellite DNA, and polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) variants. Restriction fragment length polymorphism results from the alteration of the restriction site(s) recognised by a specific restriction endonuclease or by the insertion or deletion of sequence between two restriction sites. The variation in fragment lengths is detected using gel electrophoresis. Although RFLPs were the first genetic markers developed, they are losing popularity as a screening method to identify genetic markers because they have the disadvantages of not identifying all of the polymorphism with a length of DNA, are time-consuming and restriction enzymes and consumables tend to be expensive. Simpler marker systems have subsequently been developed, many of these systems are now based on satellite DNA sequences.
Throughout the genome of higher eukaryotic organisms, there are a variety of different short DNA sequence repeats known as satellite DNA. These sequences do not code for protein and are highly variable from individual to individual in both the number and type of repeats (Groth et al., 1987). Microsatellites are composed of DNA repeats in tandem at each locus. The tandem repeats are usually simple, and consist of either a single nucleotide or dinucleotide such as (CA)n, with each dinucleotide repeated about ten times. Minisatellites have longer repeated sequences than microsatellites, such as (ACTG)n. Since microsatellites and minisatellites show a substantial amount of polymorphism, they can serve as useful markers for the identification of genetic variation of value to animal breeding. Although the variation in the number of repeats can sometimes be detected using RFLP, PCR is generally used to amplify the polymorphic region and the amplimer analysed for length variation (a technique referred to amplified fragment length polymorphism – AFLP).
PCR is also used in conjunction with SSCP. The PCR-SSCP technique offers a rapid, sensitive and relatively inexpensive way to screen for sequence variation with minimal sequencing. First described by Orita et al. (1989), this technique has become one of the preferred methods for screening samples to detect polymorphism because it is both simple and sensitive. In this techniques, regions of the gene of interest are amplified using PCR and the products denatured and then cooled rapidly to promote the formation of secondary structures due to internal base-pairing, which are in turn sequence dependent (Orita et al., 1989). The folded single-stranded DNA molecules are separated by polyacrylamide gel electrophoresis under non-denaturing conditions. The folded secondary structures are affected by physical conditions such as temperature, percentage of polyacrylamide, ionic strength of the electrophoretic buffer, glycerol concentration (Spinardi et al., 1991), ratio of acrylamide to bis-acrylamide, run length and run voltage. This can be exploited when optimising an SSCP protocol so that maximum variation can be detected in a given section of DNA. Molecules that differ by even a single nucleotide may form different conformers under a given set of conditions and, upon electrophoresis in a non-denaturing polyacrylamide gel, migrate differently. Many methods for viewing the folded DNA conformers have been described. These include the radioactive labelling of primers followed by autoradiography (Orita et al., 1989), silver staining (Sanguinetti et al., 1994), ethidium bromide staining (Yap and McGee, 1993) and more recently the use of fluorescently labelled primers and fluorescent dyes.
There are several ways to identify genetic markers, but the two approaches most commonly used are the genome scanning or linkage analysis and the candidate gene approach. In the genome scan approach, the whole genome is searched to identify Quantitative Trait Loci (QTL) that affect any given trait. These are not necessarily the genes that are responsible for trait variation, but give an indication of where such genes may lie. Linkage analysis is an involved process. A map of the chromosomes, laying out the location, phase and order of genes and markers, and the distance between them, is required before linkage analysis can be performed. Firstly, a selection of about 200 markers distributed throughout the genome are genotyped, in the sire of the animals. Only the informative markers are genotyped in the progeny and each marker tested for suggestive linkage. Regions showing suggestive linkage are then studied by saturating the region with markers to identify those that are tightly linked. Phenotypic variation is then linked to the segregation of DNA markers within a population. Once the gene locus is identified by the tightly linked markers, the DNA can be sequenced. Linkage analysis can be an expensive and lengthy process requiring access to full chromosome libraries and arrays of markers.
In the candidate gene approach, known genes or gene markers that are thought to be responsible for the phenotypic variance of a trait are targeted for investigation. In this case, knowledge of the understanding of the genes that are likely to affect wool quality. The method requires a good knowledge of the physiological and biochemical processes of the gene product and can be a more direct method than the gene mapping approach, provided the right initial assumptions are made. One of the limitations of this approach is its “hit and miss” nature. A targeted gene may not be polymorphic in a population or genetic variation within the targeted gene may not affect the trait (Goddard, 2002). For the candidate gene approach to be useful, a quick and relatively inexpensive way to screen the target gene for polymorphism is essential.
The wool fibre is a complex structure composed primarily of proteins from the keratin family, which are the keratin intermediate-filament proteins (KRTs) and the keratin intermediate-filament associated proteins (KAPs). The KRTs form the skeletal structure of the wool fibre (microfibrils) and are embedded in a matrix of KAPs (Powell and Rogers, 1986), the different proteins being connected through disulphide cross-linkages (Powell, 1996). Therefore, genes that code for the KAPs and KRTs proteins are potential candidate genes in the identification of genetic markers associated with wool quality traits.
Half-sib analysis is a tool that allows genetic effects to be ascertained from field trials while controlling for environmental and management effects. Firstly, the gene being targeted must be polymorphic, with at least two alleles. A good sire is selected and mated to many ewes (at least 200 in number), that are selected at random from a range of environments, in order to maximize phenotypic variation in wool traits. The sire must be informative at each locus that is being investigated (i.e., the genotype of the sire must be heterozygous). If not, then the progeny does not get genotyped for those loci that the selected sire is homozygous. For those loci that the sire is heterozygous, the progeny born are genotyped soon after birth, and allowed to grow until their wool measurements can be taken at (12, 24 and 36 months of age). Suppose a sire has the genotype AB at the K33 locus, then all the progeny that have inherited the A allele from the sire are put in one group, and those that have inherited the B allele from the sire are put in another group. The means of the wool measurements from both groups are then compared. If the group of progeny that inherited the B allele from their sire are found to for example have a significantly stronger staple strength than those progeny that inherited the A allele from their sire, then this would give an indication that the K33 B allele might be associated with stronger staple strength.
Numerous studies have described variation within both the KAP and KRT genes, including the work of Rogers et al. (1994a); Parsons et al. (1994a; 1996); McLaren et al. (1997); Beh et al. (2001); Itenge-Mweza et al. (2007). There are some reports associating variation in the KRT and KAP genes with variation in wool traits. Parsons et al. (1994b) and Beh et al. (2001) reported associations between variation in KAPs and mean fibre diameter in Merino sheep, while Rogers et al. (1994b) reported association between staple strength in Romney sheep and the region spanning the KAP1.1/KAP1.3/K33 loci on ovine chromosome 11. Itenge et al. (2009; 2010) reported association between variation in the KAP1.1 gene with variation in yield. In one of the half-sib families studied, variation in the K33 gene was associated with variation in staple strength. Markers, other than the KRT and KAP genes associated with wool traits have also been reported and these, together with reported keratin gene markers are summarised in Table 1.1.
Gel electrophoresis is the process in which an electrical current is applied to a gel to separate large molecules such as nucleic acids, from a mixture of similar molecules, based on differences or how they react to the electrical current. The technique relies on the fact that
Potential genetic markers for wool quality traits reported by various researchers.
nucleic acids are negatively charged because of the phosphate groups on the phosphodiester backbone of the nucleic acid strands (Nicholl, 1994). Nucleic acid molecules will migrate from the negative (black) terminal to the positive (red) terminal if put in solution and an electric field is applied, due to the net negative charge in solution. The gel matrix adds a sieving effect so that particles can be characterized by both charge and size.
Agarose is a macromolecular substance that is derived from seaweed. It can be purified to a whitish granular powder which, when mixed with water and heated, can be left to set like a jelly. This is called a gel and it acts like a sieve for the DNA molecules. To separate DNA molecules that are different lengths, agarose is used to produce a molecular sieve. The speed that the DNA travels through the gel is inversely proportional to the size of the DNA. In other words, small DNA particles migrate faster than large DNA molecules, as they are less physically restrained by the gel matrix. The length of a piece of DNA can be determined by comparing it to a molecular weight ladder. Agarose gel electrophoresis can be affected by:
The percentage of agarose, which affects the sieving of the DNA molecules.
The voltage applied during the electrophoresis, which cause the DNA molecules to move.
Typically, 1000 – 50,000 bp can be separated by 0.3% agarose, and 300 – 6000 bp can be separated by 1.4% agarose, while base pairs less than 500 are better separated using polyacrylamide gel, with gel percentage between 10-20. The polyacrylamide gel electrophoresis works under non-denaturing conditions.
After the electrophoresis is complete, the molecules in the gel can be stained to make them visible. Ethidium bromide, silver, or coomassie blue dye may be used for this process. Other methods may also be used to visualize the separation of the mixture\'s components on the gel. If the analyte molecules fluoresce under ultraviolet light, a photograph can be taken of the gel under ultraviolet lighting conditions, often using a Gel Doc. A molecular weight marker (MM) is often included on the gel to give an indication of the fragment size.
An example of a gel photo. MM is the molecular marker. Lane 1 has 341 bp DNA, while lane 4 has 280bp DNA. Lane 2 is blank.
This paper discusses the identification of genetic variation in the KAP3.2, KAP6.1, KAP7, KAP8, KRT2.10 and BfMS loci in Merino sheep using polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis. Polymorphism within these loci is likely to be in part responsible for the observed variation in wool characteristics and could result in the identification of gene markers to be used in gene marker-assisted selection programmes within the wool industry.
This study used two half-sib families referred to as Sire Line 1 (SL1) and Sire Line 2 (SL2). The SL1 half-sib was produced by mating a fine wool producer Merino ram to 150 Merino ewes, selected at random from a range of New Zealand environments, in order to maximise phenotypic variation in wool traits. In year one, the SL1 consisted of 131 pure New Zealand Merino lambs, with 128 of these surviving to the second shearing at 24 months. Following the second shearing the wether lambs and some of the ewe lambs were culled and only the remaining ewe lambs (n = 37) were shorn at 36 months of age. The SL2 half –sib consisted of 35 lambs (Merino x Romney ram x Merino ewes). Half-sib groups were kept as single flocks to minimise environmental variation between individual progeny and provide control. All lambs were tagged at birth to their dam and their gender and birth rank were recorded.
Mid-side wool samples were collected at 12, 24 and 36 months of age for SL1 and at 12 months of age for SL2. Except for greasy fleece weight (GFW) which was determined at shearing, wool measurements were performed by the New Zealand Wool Testing Authority Ltd (NZWTA), Napier, New Zealand according to International Wool Textile Organisation (IWTO) standards. Measurements included comfort factor or the percentage of fibres of diameter greater than 30 µm (F<30), mean fibre diameter (MFD, IWTO-12-03), fibre diameter standard deviation (FDSD, IWTO-12-03), coefficient of variation of fibre diameter (CVD, IWTO-12-03) and curvature, were all measured using a Sirolan™ Laserscan Fibre Diameter analyser while the mean staple length (MSL, IWTO-30) and mean staple strength (MSS, IWTO-30) of each sample was determined using Automatic tester for Length and Strength (ATLAS). The colour (MY-Z) and brightness (MB) of the wool was measured using a reflectance spectrophotometer, where the tristimulus values Y-Z indicate the yellowness of the wool and the tristiulus value Y represents the brightness of the wool. The yield of wool, the weight of clean wool after impurities such as vegetable matter have been removed, expressed as a percentage of greasy wool weight was mathematically derived for the wool base (IWTO-19) measurements. Once yield measurements were obtained from the NZWTA, clean fleece weight (CFW) was calculated as the product of GFW and yield.
Blood samples (containing DNA) were collected from the progeny and sires onto FTATM cards (Whatman, Middlesex, UK). These were stored at room temperature (See Figure 2.1). A small punch (1.2 mm in diameter) was taken from the blood on the FTATM cards using a Harris Micro Punch (Whatman International Ltd, UK) and put into a 200 μL tube. The DNA on the punches was isolated following a modified manufacturer’s protocol. 200 μL of FTATM reagent was added to each tube containing a 1.2 mm punch of FTATM paper, containing the sample DNA. The tubes were incubated at room temperature for 60 minutes. Each tube was vortexed three times for about five seconds at the start of the incubation, half-way through the incubation, and after the incubation period. The FTATM reagent was aspirated, and the cards were washed with 200 μL of TE buffer (1 M Tris and 0.5 M Na2EDTA) for two minutes. The TE buffer was aspirated and the tubes were left open, but covered with a tube holder and stored at 4 oC and used for the subsequent PCR reaction.
The PCR conditions for the loci that are described in the literature were initially used. However, re-optimisation was necessary for amplification in an i-Cycler PCR machine (Bio-Rad Laboratories Inc., Hercules, CA, USA). The PCR protocols were optimised by using a temperature gradient (to determine annealing temperature) coupled with a magnesium titration.
All the primer sequences used in the study were obtained from the literature (Table 2.1), and were synthesized by Invitrogen New Zealand Limited, Penrose, Auckland, New Zealand. PCR amplifications were performed in a reaction mixture containing ~ 50 ng of genomic DNA on a washed 1.2 mm punch of FTATM paper, 1× PCR reaction buffer with 1U Taq polymerase (Qiagen, GmBH, Hilden, Germany). Table 2.2 lists the total reaction volume used along with the specific dNTP, primer, magnesium, and Q concentrations for each locus.
Amplification consisted of 1 minute denaturation at 95 oC, followed by 30 cycles of denaturation at 95 oC for 1 minute, annealing at temperatures specified in Table 2.3 for 1 minute and extension at 72 oC for 1 minute, with a final extension of 72 oC for 7 minute. All the primer sequences used in the study were obtained from the literature, and were synthesised by Invitrogen New Zealand Limited, Penrose, Auckland, New Zealand.
FTATM cards of blood samples collected from the progeny of sire line 1
Primer sequences and source references for each locus investigated.
Locus | Total | Primer | dNTP | Mg2+ | Q |
volume (µL) | concentration (nM) | concentration (µM) | concentration (mM) | Conce- tration (×) | |
KAP3.2 | 25 | 350 | 175 | 1.0 | 1× |
KAP6.1 | 25 | 400 | 200 | 1.0 | 1× |
KAP7 | 25 | 350 | 175 | 1.0 | 1× |
KAP8 | 25 | 350 | 175 | 1.0 | 1× |
KRT2.10 | 20 | 400 | 200 | 1.0 | - |
BfMS | 25 | 350 | 175 | 1.0 | 1× |
Optimised PCR conditions for each locus investigated.
Locus | Annealing temperature (oC) | Amplimer size (bp) |
KAP3.2 | 58 | 424 |
KAP6.1 | 62 | 528 |
KAP7 | 63 | 413 |
KAP8 | 62 | 124* |
KRT2.10 | 65 | 191 |
BfMS | 58 | 200* |
Optimised annealing temperatures and predicted amplimer sizes for each locus investigated.
Amplimers were analysed in 1.0% w/v SeaKem® LE agarose (FMC Bioproducts, Rockland, Maine, USA) gels prepared with 1× TBE buffer (89 mMTris, 89 mM orthoboric acid, 2 mM Na2EDTA; pH 8) containing 0.1 mg/L ethidium bromide. Five µL of PCR product was added to 2.5 µL of loading dye (0.2% bromophenol blue, 0.2% xylene cyanol, 40% (w/v) sucrose) and the gels were electrophoresed at a constant 10 Vcm-1 for 30 minutes. A molecular weight marker (Invitrogen Life Technologies) was included on the gel to give an indication of the fragment size. DNA bands were viewed on a UV transilluminator (254 nm) and a photograph taken for records.
PCR-SSCP conditions were available in the literature for KAP3.2 (McLaren et al., 1997), however these were deemed to be insufficiently stringent. For this reason, the PCR-SSCP protocols used in this study were established empirically using template DNA from two small half-sib families (to observe inheritance of allele-specific banding pattern) and DNA samples of other unrelated Merino sheep (for increased genotypic variation). Many different gel conditions (gel percentage, voltage, time of running, temperature, addition of glycerol) were assessed to determine the optimum combination of conditions to resolve allele specific banding patterns in a reproducible manner. Amplimers from sires of the SL1 and SL2 and their selected progeny were also included on the optimising gels in order to ascertain allele banding patterns by following inheritance, and to determine whether the sires were heterozygous, and therefore informative, for the locus genotyped. Alleles were named in the order they were identified using letters of the alphabet.
Each locus used specific SSCP gel conditions, and these are summarised in Table 2.4. Polyacrylamide (37.5:1 acrylamide / bis-acrylamide, Bio-Rad Laboratories, Hercules, Ca, USA) vertical gels (Protean II 16 x 16 cm, 1.0 mm thick spacers, 28 well comb, Hoefer, Inc., San Francisco, Ca, USA) were prepared containing 0.5× TBE (44.5 mMTris, 44.5 mM orthoboric acid, 1 mM Na2EDTA [pH 8.0]) and polymerised using 10% ammonium persulphate and TEMED. Gels were pre-electrophoresed at running temperatures and voltage for one hour. Amplimers were mixed with 50 µL loading dye (95% formamide, 10 mM Na2EDTA, 0.025% bromophenol blue, 0.025% xylene cyanol), denatured by heating at 95 oC for five minutes and immediately placed on wet ice before loading 15 µL aliquots. The gels were then electrophoresed at the optimum gel conditions with 0.5× TBE running buffer, followed by silver-staining according to the method of Sanguinetti et al. (1994).
For KAP3.2, KAP7 and KRT2.10 loci, genomic DNA was obtained from the sire and this DNA was amplified using the PCR conditions described above and the amplimers were subsequently cloned using the Promega pGEM® - T Easy Vector System I (Promega Corporation, Madison, WI, USA). Since each plasmid can only accept one molecule of DNA and therefore only one allele. Ligation reactions were performed in a total reaction volume of 10 μL containing three units T4 DNA ligase, 50 ng of plasmid DNA and 1× ligation buffer, and incubated overnight at 4 oC. Constructs were transformed into competent E. coli cells (InvitrogenTM, One ShotTM, INVαF’) using the manufacturer’s protocol. Sixty μL and 150 μL of the transformation mix were spread on labelled LB (0.5 % casein hydrolysate, 0.25 % yeast extract, 85.6 mM NaCl; pH 7.0) agar plates containing 100 μg/mL ampicillin that had been spread with 40 μL of 40 mg/mL X-Gal (BDH Laboratory Suppliers, Poole, England). The plates were incubated overnight at 37 oC. Six colonies for each representative allele were selected and cultured overnight in terrific broth (Invitrogen Corporation, Paisley, Scotland, UK), supplemented with 50 μg/mL ampicillin, for plasmid isolation. Colonies were screened for the correct alleles using a rapid boiling-PCR method, where by Fifty µL aliquot of an overnight culture (bacterial cells with gene of interest cultured in terrific broth) was centrifuged at 13,000 rpm for 2 minutes, the supernatant was discarded and 30 μL TE (1 M Tris, 0.5 M Na2EDTA) buffer added, boiled for 10 minutes, centrifuged at 13000 rpm for 2 minutes and then 1µL of the supernatant was used as the template for the appropriate PCR. Amplimers were run on 2% agarose gels next to the original genomic PCR amplimers for comparison. Plasmid DNA was then isolated from clones, which had banding patterns corresponding to the original banding pattern seen from amplimers of genomic DNA, using the FastPlasmidTM Mini Kits (Eppendorf, Hamburg, Germany) following the manufacturer’s instructions. These amplified plasmid DNAs were subsequently sequenced and used as standards for scoring unknown genotypes.
Locus | Gel % (37.5:1)1 | Run length (hours) | Temperature2 (oC) | Voltage (V) |
KAP3.2 | 8 | 17 | 30 | 250 |
KAP7 | 10 | 4 | 20 | 200 |
KAP8 | 10 | 4 | 20 | 200 |
BfMS | 12 | 7 | 15 | 300 |
Optimised SSCP conditions for the loci investigated.
Plasmid standards were sequenced in the forward and reverse directions using the M13 forward and reverse primers at the Waikato University DNA Sequencing Facility, University of Waikato, New Zealand or Lincoln University Sequencing Facility, Lincoln, New Zealand. The sequences were compiled using DNAMANTM version 4.0 (Lynnon Biosoft, Quebec, Canada) and the electropherograms. To minimise the likelihood of PCR and sequencing errors, sequence data was derived from four separate colonies, at least two of which were from independent PCR amplifications. When sequencing data was consistent, the sequences were submitted to NCBI GenBank (http://www.ncbi.nlm.hih.gov). These were Ovis aries keratin intermediate-filament Type II (KRT2.10) gene: Accession number AY437406; Ovis aries high-sulphur keratin IF-associated protein 3.2 (KAP3.2) gene: Accession number AY483216 and Ovis aries high-glycine/tyrosine Type II keratin protein 6.1 (KAP6.1) gene: Accession number AY483217.
In order for any of the loci to be informative, they have to be heterozygous in the chosen sires allowing the segregation of the sire alleles to be followed in the progeny and segregation analyses performed. Segregation of the sire alleles within SL2 was observed and a chi-square goodness of fit test performed to ascertain whether the sire alleles inherited by the progeny occurred in a 1:1 ratio within the population. Any progeny which had the same genotype as both its sire and dam was excluded from the association analysis since it was not possible to determine which of the alleles had been inherited from the sire. The association of alleles of KAP8 with all measured wool traits (MFD, FDSD, CVD, curvature, yield, yellowness, brightness, comfort factor, staple length, staple strength, GFW and CFW) was then analysed for each year of phenotypic data using an analysis of variance (ANOVA) tests using SPSS version 13 (SPSS Science Inc., Chicago, IL, USA). The ANOVA model included sire allele and gender as factors and a full factorial model was used. The analysis used assumed that the ewe’s alleles effects were distributed randomly in progeny. The date of birth was not included in the ANOVA because the progeny were half-sibs born in a five weeks period, and it was assumed that variation in birth date was balanced across the half-sib in the segregation analyses, and that none of the genes analysed had a significant effect on gestation length.
Six loci (KAP3.2, KAP6.1, KAP7, KAP8, KRT2.10 and BfMS) were included in the study. All of them were amplified successfully using PCR and polymorphism was detected in three loci (KAP3.2, KAP8 and BfMS). Of the loci which were polymorphic, only KAP8 was heterozygous for SL2 (Tables 3.1), and thus potentially informative as a genetic marker. The remaining loci appeared to be homozygous in the sires, and thus uninformative. Table 3.2 shows the genotype of SL2 progeny at KAP8 locus.
Locus | No. of alleles detected | SL1 genotype | SL2 genotype | Informative1 (Yes / No) |
KAP3.2 | 3 | AA | AA | No |
KAP6.1 | 1 | AA | AA | No |
KAP7 | 1 | AA | AA | No |
KAP8 | 4 | AA | AB | Yes2 |
KRT2.10 | 1 | AA | AA | No |
BfMS | 3 | AA | CC | No |
Genotype results for the loci investigated in the study, indicating whether the sire genotype was informative (heterozygous) or non-informative (homozygous).
Lamb identity | Ewe identity | Lamb genotype |
1027 | 86 | AB |
1028 | 162 | AA |
1029 | 57 | BB |
1030 | 114 | AA |
1031 | 59 | AB |
1032 | 59 | AA |
1033 | 105 | AA |
1034 | 89 | AA |
1035 | 51 | AB |
1036 | 49 | AD |
1037 | 120 | AA |
1038 | 56 | AB |
1039 | 65 | AC |
1040 | 47 | AB |
1041 | 119 | BB |
1045 | 77 | BB |
1046 | 155 | AA |
1048 | 14 | BB |
1049 | 130 | AA |
1050 | 161 | AA |
1051 | 84 | BB |
1052 | 150 | BB |
1053 | 17 | BB |
1054 | NT | AA |
1055 | 113 | AB |
1056 | 140 | BB |
1057 | 21 | AB |
1058 | 87 | AA |
1059 | 135 | AB |
1060 | 137 | AA |
1061 | 117 | AC |
1062 | 38 | AB |
1063 | 148 | AB |
1064 | 68 | AB |
1065 | . | ? |
1067 | 116 | AA |
1068 | 58 | ? |
1069 | 64 | AA |
Genotype of KAP8 SL2 progeny
Four banding patterns were identified for the KAP8 microsatellite amplimer using PCR- SSCP typing methods, and these were named A, B, C and D (Figure 3.1). The alleles were not sequenced. Mendelian inheritance was observed in SL2 half-sib family for KAP8 (Table 3.2). A Chi-square goodness of fit analysis to test whether the segregation of the sire alleles differed from a 1:1 ratio confirmed normal Mendelian segregation (Table 3.3).
PCR-SSCP of the 124 bp amplimer of the KAP8 microsatellite showing the four alleles identified. Amplimers were electrophoresed on a 10% non-denaturing acrylamide/bis-acrylamide gel for 4 hours, 200 V at room temperature (20 oC). Genotype of an individual animal is shown below each lane. SL2 genotype (AB) is bolded, and his randomly selected half-sib progeny are shown in italics.
SL2 Genotype | AB |
Number of progeny inheriting allele A | 17 |
Number of progeny inheriting allele B | 12 |
Number of progeny genotyped same as the sire | 5 |
Total number (n) | 34 |
χ2 | 0.8621 |
P-value1 | 0.3532 |
Segregation of sire alleles within SL2 sire-line. Chi-square goodness of fit was used for to ascertain whether the sire alleles inherited by the progeny occurred in a 1:1 ratio within the population. Probability values (P values) are given.1A P-value > 0.05 means that the allele segregation did not differ significantly from a 1:1 ratio.
SL1 was homozygous at the KAP8 locus based on SSCP gel patterns, and hence uninformative. SL2 was heterozygous at the KAP8 locus, having the genotype AB. Eleven out of 36 progeny had the genotype AB (Table 3.2), which was the same as that of the sire. The genotype of the ewes for these lambs was subsequently determined. Five of the ewes genotyped as AB, and the progeny of these ewes were excluded from further statistical analysis as the allelic contribution from the sire could not be determined.
The sire alleles at the KAP8 locus showed a Mendelian pattern of inheritance and segregated in a 1:1 ratio in the progeny of each half sib (Table 3.3). Statistical analyses within sire SL2 half-sib family showed that there were no association between the sire alleles (or gender) and variation of wool traits.
The number of differences between alleles within sire-lines which were not statistically significant suggested the possibility of Type II errors (failing to detect a difference when in fact there is one). To address this issue, a power analysis was conducted for each trait within each of the sire-lines to determine whether the sample sizes available were adequate to detect at least 10% differences between alleles, within each sire-line, at P<0.05 with 80% power, i.e. nper allele= (8 × 2 × Error Mean Squareestimate)/(0.1 × trait averageacross sire-lines)2.
This equation was then rearranged to allowed the actual detectable difference to be calculated for each sire-line, i.e. % detectable difference = [
KAP3.2 and BfMS were found to be polymorphic in the progeny used in this study, although they appeared to be homozygous for both sires used (Figures 3.2 and 3.3, respectively). This was confirmed with cloning and sequencing amplimers derived from sire SL1.
Sire-line | Nlambs | Trait1 | Trait average2 | EMS3 | N per allele to detect at least a 10% difference | N lambs required to detect a 10% difference |
SL2 | 29 | Prickle factor | 1.61 | 1.75 | 1080 | 2159 |
29 | MFD | 19.07 | 10.57 | 46 | 93 | |
29 | FDSD | 3.89 | 67.23 | 7103 | 14207 | |
29 | CVD | 20.46 | 91.34 | 349 | 698 | |
29 | Curvature | 94.58 | 0.45 | 0 | 0 | |
29 | Yield | 71.84 | 9.79 | 3 | 6 | |
29 | Staple length | 73.33 | 2.04 | 1 | 1 | |
29 | Staple strength | 31.30 | 12.58 | 21 | 41 | |
29 | Brightness | 69.92 | 0.27 | 0 | 0 | |
29 | Yellowness | -2.83 | 135.71 | 27195 | 54389 |
Sample size required to detect at least a 10% difference between KAP8 sire allele groups in the wool traits list for each sire-line, at P<0.05 with 80% power.
Sire-line | Nlambs | Age (months) | Trait1 | Trait average2 | Smallest detectable difference (%)3 | Difference observed between alleles (%) |
SL2 | 29 | 12 | Prickle factor | 1.77 | 86.3 | 29.5 |
29 | 12 | MFD | 19.21 | 17.9 | 3.2 | |
29 | 12 | FDSD | 3.91 | 221.3 | 1.9 | |
29 | 12 | CVD | 20.45 | 49.1 | -0.4 | |
29 | 12 | Curvature | 97.04 | 0.7 | 6.4 | |
29 | 12 | Yield | 72.69 | 4.6 | 1.1 | |
29 | 12 | Staple length | 33.56 | 2.0 | 4.3 | |
29 | 12 | Staple strength | 69.06 | 11.9 | 22.6 | |
29 | 12 | Brightness | -2.97 | 0.8 | -2.1 | |
29 | 12 | Yellowness | 72.60 | -433.1 | 7.1 |
A comparison of the smallest detectable difference between KAP8 sire-allele groups with the progeny numbers used and the observed difference between the sire-allele groups means for each wool trait measured.
Polymorphism could not be detected at the KAP6.1, KAP7 and KRT2.10 loci in any of the animals used in this study. KAP 7 was sequenced, and nucleotide sequences from SL1 KAP7 amplimer (GenBank accession number AY791846) was aligned with the published KAP7 gene by Kuczek and Rogers (1987); GenBank accession number X05638) which shows two unique sequences (Figure 3.4).
PCR-SSCP analysis of the 424 bp amplimer of the KAP3.2 gene showing the three alleles identified (A, B and C). The genotype of an individual animal is shown below each lane.
PCR-SSCP analysis of the 200 bp amplimer of the BfMS microsatellite showing the three alleles identified (A, B and C) using a half-sib test family. The genotype of an individual animal is shown below each lane. Sires SL1 and SL2 genotypes are bolded.
Alignment of the KAP7 gene sequence cloned from sire MV144-58-00 (Accession number AY791846) with Kuczek and Rogers (1987) published KAP7 gene (Accession number X05638). Upstream and downstream primers are underlined and the start and stop codons are bolded). Dashes represent same nucleotides to the nucleotide above and dots represent nucleotides missing in the other sequence.
Sex Average (cM) | Female (cM) | Male (cM) | Locus code | Marker | Marker description or associated gene |
101.9 | 84.3 | 119.3 | \\BM4129 | Sequence – tagged site | |
104.1 | 86.6 | 122.0 | \\UCDO31 | RAPD Marker | |
107.1 | 89.0 | 125.2 | \\MCM58 | Microsatellite | |
111.3 | 91.7 | 130.5 | \\BL41 | VANGL1 | |
111.3 | 91.7 | 132.2 | \\BM723 | STS | |
111.3 | 91.7 | 132.2 | \\BM723 | STS | |
113.8 | 92.9 | 133.9 | \\OARAE57 | Microsatellite | |
123.4 | 105.2 | 142.2 | \\MCMA6 | ? | |
124.5 | 106.8 | 142.2 | \\MCMA6L | ? | |
124.5 | 106.8 | 142.2 | \\BMS482 | Sequence – tagged site | |
124.5 | 106.8 | 142.2 | \\CSSM054 | Phosphoglycerate dehydrogenase | |
126.0 | 107.9 | 143.2 | PRPF3 protein | ||
126.0 | 108.9 | 143.2 | Aryl hydrocarbon receptor nuclear translocator | ||
126.0 | 108.9 | 143.2 | Trichohyalin | ||
127.3 | 111.1 | 144.3 | \\RM065 | Dinucleotide repeat | |
132.0 | 115.3 | 149.1 | ~CSAP033E | Microsatellite | |
134.9 | 120.0 | 150.4 | Immunoglobulin superfamily 9 | ||
135.7 | 121.5 | 150.4 | ATPase | ||
137.0 | 121.5 | 152.9 | ADAM metallopeptidase | ||
139.8 | 122.7 | 156.8 | \\URB006 | Sequence – tagged site | |
143.6 | 127.6 | 160.4 | \\BM6438 | Sequence – tagged site | |
143.6 | 127.6 | 160.4 | Oligodendrocyte transcription factor 2 | ||
144.8 | 127.6 | 162.2 | \\SRCRS23H | ? | |
144.8 | 127.6 | 162.2 | \\TGLA49 | Microsatellite | |
144.8 | 127.6 | 162.2 | \\DVEPC88 | Neu associated kinase | |
145.3 | 127.6 | 163.1 | Keratin associated protein 7.1 | ||
145.3 | 127.6 | 163.1 | Keratin associated protein 7.1 | ||
145.3 | 127.6 | 163.1 | Keratin associated protein 8.1 | ||
145.3 | 127.6 | 163.1 | Keratin associated protein 7.1 | ||
145.3 | 127.6 | 163.1 | Keratin associated protein 11.1 | ||
145.4 | 127.6 | 163.2 | Keratin associated protein 6.1 | ||
145.8 | 127.6 | 164.0 | Glutamate receptor, ionotropic, kainite 1 | ||
149.6 | 131.5 | 168.1 | Amyloid beta (A4) precursor protein | ||
150.5 | 131.5 | 169.5 | \\BMS574 | Sequence – tagged site | |
150.5 | 131.5 | 170.2 | \\DVEPC117 | Sequence – tagged site | |
150.5 | 131.5 | 170.2 | \\DVEPC117 | Sequence – tagged site | |
152.1 | 132.8 | 171.3 | \\BMS2321 | Sequence – tagged site | |
153.2 | 132.8 | 173.1 | \\DVEPC128 | Neural cell adhesion molecule 2 | |
157.1 | 138.0 | 176.7 | \\RM095 | Dinucleotide repeat | |
158.1 | 138.0 | 177.6 | \\MAF64 | Dinucleotide repeat | |
169.2 | 150.1 | 188.2 | \\ILSTS004 | Sequence – tagged site | |
171.1 | 152.6 | 188.2 | \\DVEPC54 | Microsatellite | |
174.4 | 154.8 | 194.2 | \\MCMA8 | Sequence – tagged site | |
176.0 | 154.8 | 197.2 | \\MNS94 | Microsatellite | |
193.1 | 169.9 | 216.0 | \\CSSM004 | Microsatellite | |
195.3 | 171.1 | 219.6 | \\BMS4000 | Sequence – tagged site | |
200.0 | 177.1 | 223.6 | \\UCDO46 | ? |
Linkage map for part of ovine chromosome 1 (modified from http://rubens.its.unimelb.edu.au/~jillm/jill.htm). The bolded genetic markers were investigated in this study.
Sex Average (cM) | Female (cM) | Male (cM) | Locus Code | Marker | Marker description or associated gene |
149.6 | 151.4 | 148.8 | \\BMS695 | Sequence – tagged site | |
149.6 | 151.4 | 148.8 | \\BM827 | Microsatellite | |
152.5 | 151.4 | 153.4 | \\MCM141 | ? | |
153.1 | 151.4 | 154.3 | \\OARSHP2 | Microsatellite | |
153.1 | 151.4 | 154.3 | \\ILSTS042 | Sequence – tagged site | |
154.1 | 151.4 | 156.3 | \\BMS424 | Sequence – tagged site | |
163.1 | 155.2 | 170.4 | \\BP1 | Blood pressure QTL1 | |
163.1 | 155.2 | 170.4 | \\DU469297 | ? | |
165.6 | 160.5 | 170.4 | \\EPCDV025 | ? | |
167.1 | 166.2 | 170.4 | KIT Ligand | ||
168.7 | 166.6 | 172.4 | \\UCDO13 | ? | |
170.7 | 166.6 | 174.3 | Keratocan | ||
170.7 | 166.6 | 174.3 | Lumican | ||
177.8 | 172.1 | 182.4 | \\AGLA293 | Microsatellite | |
179.4 | 174.9 | 183.5 | ~CSAP017E | Microsatellite | |
179.4 | 174.9 | 183.5 | \\OARFCB5 | Dinucleotide repeat | |
179.4 | 174.9 | 183.5 | Glycosylation dependant cell adhesion molecule | ||
179.4 | 174.9 | 183.5 | \\OARHH38 | Microsatellite | |
180.0 | 176.2 | 183.5 | \\ILSTS022 | Sequence – tagged site | |
180.0 | 176.2 | 183.5 | Retinoic acid receptor 8 | ||
182.9 | 178.7 | 186.5 | Keratin | ||
183.9 | 181.2 | 186.5 | Keratin | ||
183.9 | 181.2 | 186.5 | \\BMC1009 | Similar to intermediate filament type II keratin | |
186.3 | 181.2 | 190.9 | \\CABB011 | Genomic survey sequence | |
188.2 | 185.3 | 190.9 | \\CSSM034 | Microsatellite | |
188.2 | 185.3 | 190.9 | Histone deacetylase 7A | ||
188.2 | 185.3 | 190.9 | \\UCDO52 | ? | |
195.5 | 188.6 | 201.0 | \\BL4 | Bell-like homeodomain protein 4 | |
197.0 | 190.4 | 202.8 | Lysozyme | ||
198.6 | 191.8 | 204.6 | \\CSRD2125 | ? | |
199.1 | 191.8 | 205.5 | Interferon gamma | ||
199.1 | 191.8 | 205.5 | Interferon gamma | ||
199.1 | 191.8 | 205.5 | Interferon gamma | ||
202.2 | 195.4 | 207.8 | \\BMS1617 | STS | |
204.1 | 196.7 | 210.6 | \\OARVH34 | Microsatellite | |
206.2 | 197.7 | 213.7 | \\BR2936 | Sequence – tagged site | |
207.0 | 197.7 | 215.5 | \\OARVH130 | Microsatellite | |
207.0 | 197.7 | 215.5 | \\MAF23 | Microsatellite | |
209.0 | 199.0 | 218.1 | \\OARCP43 | Microsatellite | |
214.7 | 208.6 | 219.8 | \\RM154 | Tandem repeat region | |
218.5 | 211.3 | 223.7 | Insulin like growth factor | ||
218.5 | 211.3 | 223.7 | Insulin like growth factor | ||
218.5 | 211.3 | 223.7 | Insulin like growth factor | ||
218.5 | 211.3 | 223.7 | Insulin like growth factor | ||
223.9 | 215.4 | 231.6 | \\CSRD2111 | ? | |
224.4 | 215.4 | 232.5 | ~CSAP009E | ? |
Linkage map for part of ovine chromosome 3 (modified from http://rubens.its.unimelb.edu.au/~jillm/jill.htm). The bolded genetic markers were investigated in this study.
Four alleles, designated A, B, C and D were identified at the KAP8 (CA)n repeat microsatellite locus using PCR-SSCP in this study. The microsatellite at the KAP8 locus was included in the study because this region is highly polymorphic, with 15 alleles previously reported (Wood et al., 1992) using denaturing polyacrylamide gel electrophoresis, while Parsons et al. (1994a) detected four allelic fragments (123, 125, 133 and 139 bp) at the same locus using the methods by Wood et al. (1992) in a Merino half-sib family. Only SL2 was heterozygous at the KAP8 microsatellite in this study. SL1 was homozygous, despite the reported highly polymorphism in this locus (Wood et al., 1992). The method used to detect polymorphism in this study differed to that of (Wood et al., 1992), which used denaturing polyacrylamide gel electrophoresis. In this study, PCR-SSCP was used because this technique is simple, sensitive, relatively inexpensive and routinely used in the laboratory where the research was carried out. It is possible that if the original technique was employed, more alleles may have been observed at this locus.
Neither of the SL2 alleles were associated with variation in the wool traits that were measured (data not shown). The possibility of this locus having an affect on wool traits cannot be ruled out however, because only two alleles (that were the genotype of SL2) were analyzed, and that the sample numbers used in the study were relatively small (n = 29). Power analysis results (Table 9.5) showed that the observed differences between the sire allele groups were smaller than the smallest detectable difference for MFD, FDSD, CVD, curvature, yield, staple length, brightness and yellowness and therefore the possibility of making a Type II error (i.e. not detecting an association when there was one) is likely. Variation in MFD has previously been significantly associated with alleles at the KAP8 locus (Parson et al., 1994a). The authors did not describe the alleles associated, and no sequence data was presented. Though alleles at the KAP8 microsatellite locus were not sequenced in this study, it is possible that SL2’s alleles were different from those associated with differences in average MFD by Parsons et al., (1994a).
Three alleles, designated A, B and C were identified at the KAP3.2 locus. However, both sire lines were homozygous, and thus uninformative. McLaren et al. (1997) identified two alleles at the KAP3.2 locus using PCR-SSCP methods. KAP3.2 (together with KAP1.1, KAP1.3 and K33) have been mapped to ovine chromosome 1 (Figure 3.5). Variations in all of the three genes (KAP1.1, KAP1.3 and K33) have been previously associated with variation in wool traits (Itenge et al., 2009; Itenge et al., 2010; Rogers et al., 1994b). It is therefore suggested that sires that are heterozygous get investigated in further studies. Three alleles, designated A, B and C were identified at the BfMS microsatellite. Bot et al. (2003) reported eight alleles at the BfMS locus. Two of these alleles were significantly associated with CFW and GFW. However, both sire lines were homozygous, and thus uninformative at the BfMS locus.
Polymorphism could not be detected at the KAP6.1, KAP7 and KRT2.10 loci in this study, although all of these genes have been reported to be polymorphic in the literature (Parsons et al., 1993; McLaren et al., 1997). The reported polymorphism in KRT2.10 (two alleles) and KAP7 (four alleles) was identified using PCR-RFLP (McLaren et al., 1997) whereas the polymorphism within KAP6.1 (two alleles) was revealed with PCR-SSCP of AluI-digested PCR amplimers (McLaren et al., 1997). Parsons et al. (1993) reported a diallelic polymorphism using BamHI PCR-RFLP to give alleles designated A1 (24.5 kb) and A2 (14.1 kb). However, no sequence data was presented. Since only two KAP6.1 alleles have previously been reported, thus it was accepted that SL1 was homozygous at this locus without further sequencing although this locus could still be polymorphic which only sequencing would reveal. The KAP6.1 amplimers were also subjected to a variety of PCR-SSCP conditions in an effort to detect sequence variation. Digestion of the amplimer with AluI or BamHI as per McLaren et al. (1997) was not performed, however, and it is possible that this may have revealed variation at KAP6.1. KRT2.10 has been mapped to ovine chromosome three and two alleles have been reported at the KRT2.10 locus using a BsrDI PCR-RFLP (McLaren et al, 1997). Genes coding for the KRT proteins are highly conserved during evolution (Powell, 1996; Marshall and Gillespie, 1982), and do not have much variation within them. Therefore, it was easy to accept that the KRT2.10 locus (with only two alleles) was likely to be homozygous. The fact that the KRT proteins are highly conserved during evolution (Powell, 1996; Marshall and Gillespie, 1982) suggests that genes coding for these proteins are intolerant to major changes and that they are very important to the integrity of the wool fibre.
Loci that were polymorphic, but uninformative in this study (KAP3.2, BFMS) )need to be investigated further. Sires that are heterozygous at these loci need to be identified and used in half-sib analysis. Other loci that map to the same chromosome regions as the keratin genes investigated in this study are also worth of investigating in the future as potential gene markers for wool quality traits. On chromosome 1, future genes of interest include KAP11.1 and genes coding for trichohyalin (a very important wool follicle protein) (refer to Figure 3.5). On chromosome 3, loci of interest include KRT2.13, BMC1009 (Similar to intermediate filament type II keratin), RARG (Retinoic acid receptor 8) and IGF1 (insulin like growth factor) (refer to Figure 3.6). It is worth noting that previous studies by Damak et al. (1996) have shown positive effects of IGF1 on wool traits. Transgenic sheep produced by pronuclear microinjection with a mouse ultra-high-sulphur keratin promoter linked to an ovine IGF1 resulted in significant increase of CFW and bulk in transgenic sheep compared to non-transgenics, although MFD did not show significant differences (Damak et al.,1996).
There are other genes that have not been positioned on the linkage map that may be potential gene markers for wool quality traits. Some of these have already been associated with wool quality traits. These include the retinoic acid receptor (RARα) (Nadeau et al., 1992), homeobox proteins (HOX2) (Nadeau et al., 1992) and growth hormone (Hediger et al., 1990). Retinoic acid induces expression of genes such as homeobox and KRTs and there is a possibility that retinoic acid is involved in the regulation of KAPs, given its genomic position on chromosome 11 (Parsons et al, 1994c). Growth hormone has been positioned on chromosome 11 through in situ hybridization (Hediger et al., 1992). Furthermore, there have been numerous reports with variable effects of growth hormone on wool characteristics. For example, Ferguson (1954) and Johnson et al. (1985) observed significant increase in GFW during the injections of growth hormone. In contrast, no effect of recombinant growth hormone on CFW was found in a study by Zainur et al. (1989). Wheatley et al. (1966) found that growth hormone suppressed wool growth and that there was accelerated wool growth after withdrawal of growth hormone. Polymorphism at the genes encoding growth hormone have been reported (Valinsky et al., 1990; Wallis et al., 1998; Sami et al., 1999), and different alleles of growth hormone may affect wool growth in different ways.
The search for genetic markers affecting wool quality traits is very different to genetic engineering (GE) and transgenesis. While GE involves the manipulation or modification of genetic composition of an organism, and transgenesis requires the development and use of transgenic animals, the former detects changes within the genetic make-up of an organism, but does not alter it. Marker-assisted selection may therefore be better preferred within the wider “non-scientific” community, than the use of transgenic sheep to produce superior wool traits. Transgenesis in sheep is also still in its infancy, and successful transgenesis rates are very low (less than 13%) (Powell et al., 1994). This makes marker-assisted-selection a more efficient, relatively cheaper and easier technique to improve wool quality traits than sheep transgenesis. The debate on GE will most likely continue and intensify especially where animals are involved. However, marker-assisted technology in livestock offers a powerful "green" alternative to gene manipulation.
Genetic markers are not affected by environmental noise and would allow sheep breeders to select animals with improved wool characteristics at an early age and cull the non-desirable lambs. This would speed up the process of genetic selection and decrease the generation interval. There is therefore a potential to select superior animals very early in life and not have to wait for an animal to reach its adult life to demonstrate that it has superior wool quality. This has the advantage of overcoming the limitation of “blind” selection, and increase the accuracy and efficiency of selection and result in a more profitable wool industry with direct benefits of cost to the consumer.
I thank the Almighty God for everything in my life. I wish to thank my Honours supervisor, Prof. Jim Reynoldson from Murdoch University and my PhD supervisors (Prof. Jon G. H. Hickford from Lincoln University and Dr. Rachel Forrest from Eastern Institute of Technology). I am very grateful for the AUS-AID and NZAID scholarships that I received from the Australian and New Zealand governments, respectively. I am also very grateful for the Staff Development Fellowship award that I received from the University of Namibia.
In England Anne Longfield, England’s Children’s Commissioner, has written to the biggest social media companies, urging them to commit to tackling issues of disturbing content. Her letter follows the suicide of 14-year-old Molly Russell, who tragically killed herself after viewing distressing self-harm images on Instagram. The letter urges social media companies to back the introduction of a statutory duty of care where they would have to prioritise the safety and wellbeing of children using their platforms. Ms. Longfield’s letter ends with the following message to the digital industry:
\n\n \nWith great power comes great responsibility and it is your responsibility to support measures that give children the information and tools they need growing up in this digital world—or to admit that you cannot control what anyone sees on your platforms.\n
According to literature use of the internet has risen rapidly in the last decade [1]. The way in which young people interact has changed significantly over the last decade. Social media enables them to develop online connections with people within their immediate friendship group but also to form connects with people who are more geographically dispersed. As a result of the digital revolution in recent years, young people are now able to communicate with others more efficiently and gain access to knowledge and advice more rapidly. For those living in rural communities, social media can facilitate social communications which otherwise would not be possible.
\nMy own discussions with young people in schools indicates that social media is an extremely important part of their daily lives. It brings many benefits but is also exposes them to risks. Young people are often very aware of these risks and understand how to keep themselves safe. However, sadly this does not prevent all of them from harm, as is evident through recent cases of teenage suicides as a result of social media, which have been highlighted in the media in the United Kingdom (UK) and more widely.
\nThis chapter highlights some of the detrimental and positive effects of social media use on children and young people’s mental health. The implications for schools, parents, social media and advertising companies and the government are addressed. This chapter highlights that schools cannot solve all of the problems and that other stakeholders also have a responsibility to keep young people safe when they are online.
\nResearch suggests that social media use is far more prevalent among young people than older generations [1]. Young people aged 16–24 are the most active social media users with 91% using the internet for social media [1]. Young people use social media for a variety of purposes, including for entertainment, to share information and network with others and to gain support and health information [1].
\nEvidence suggests that social media use can result in young people developing conditions including anxiety, stress and depression [1]. There are various reasons for this, and this section will explore the contributing factors. Research has found that four of the five most used social media platforms make young people’s feelings of anxiety worse [1]. Research suggests that young people who use social media heavily, i.e., those who spend more than 2 hours per day on social networking sites are more likely to report poor mental health, including psychological distress [2].
\nCyber-bullying is a significant problem which affects young people. Evidence suggests that seven in 10 young people experience cyberbullying [1]. Cyberbullying exists in a variety of forms. It can include the posting of hurtful comments online, threats and intimidation towards others in the online space and posting photographs or videos that are intended to cause distress. This is not an exhaustive list. Cyberbullying is fundamentally different to bullying which takes place in person. The victim of the bullying may find it difficult to escape from because it exists within the victim’s personal and private spaces such as their homes and bedrooms. Additionally, the number of people witnessing the bullying can be extremely large because of the potential of social media for online posts to be shared across hundreds, thousands and millions of people. For the victim this can be significantly humiliating and result in a loss of confidence and self-worth. Humiliating messages, photographs and videos can be stored permanently online, resulting in the victim repeatedly experiencing the bullying every time they go online. Victims of cyberbullying can experience depression, anxiety, loss of sleep, self-harm and feelings of loneliness [3].
\nSocial media has also been associated with body image concerns. Research indicates that when young girls and women in their teens and early twenties view Facebook for only a short period of time, body image concerns are higher compared to non-users [4]. Young people view images of “ideal” bodies and start to make comparisons with their own bodies. This can result in low body-esteem, particularly if young people feel that their own bodies do not compare favourably to the “perfect” bodies they see online. Young people are heavily influenced by celebrities and may desire to look like them. If they feel that this is unattainable it can result in depression, body-surveillance and low body-confidence. Young people can then start to develop conditions such as eating disorders. The issue of body image is not just a female issue. Young males are also vulnerable and influenced by the muscular, well-toned bodies that they see online. We now live in an age when males are taking increasing interest in their appearance and viewing images of muscular, toned bodies can result in them putting their bodies through extensive fitness regimes and males are also vulnerable to developing eating disorders.
\nThe opportunity for people to use digital editing software to edit their appearance on photographs can also result in young people developing a false sense of beauty. It is worrying that there is a rise in the number of young people seeking to obtain cosmetic surgery [1] and the popularity of “selfies” in recent years has resulted in an increase in images which portray beauty and perfection. These images can have a negative impact on body-esteem and body-confidence.
\nResearch demonstrates that increased social media use has a significant association with poor sleep quality in young people [5]. It seems that young people enjoy being constantly connected to the online world. They develop a “Fear of Missing Out” (FoMO) which is associated with lower mood and lower life satisfaction [6]. This can result in young people constantly checking their devices for messages, even during the night, resulting in broken sleep. Sleep is particularly important during adolescence and broken sleep can result in exhaustion and lack of opportunity for the brain to become refreshed. Lack of sleep quality can have a range of detrimental effects, but it can also impact on school performance and their behaviour. My own conversations with school leaders suggest that many adolescents demonstrate signs of tiredness during the school day. This can result in disengagement in lessons, thus having a detrimental effect on academic attainment.
\nThe link between social media use, self-harm and even suicide is particularly worrying [1]. The fact that young people can access distressing content online that promotes self-harm and suicide is a significant cause for concern. This content attempts to “normalise” self-harm and suicide and can result in young people replicating the actions that they are exposed to.
\nResearch suggests that young people are increasingly using social media to gain emotional support to prevent and address mental health issues [7]. This is particularly pertinent for young people who represent minority groups, including those who identify as lesbian, gay, bisexual or transgender (LGBT), those with disabilities and those representing black and minority ethnic groups. The use of social media to form online digital communities with others who share similar characteristics can be extremely powerful. Young people from minority groups are able to become “global citizens,” thus reducing isolation. Participating in online networks presents them with an opportunity to meet with others who share their identities, to gain mutual support and advice and to gain solidarity. These networks can reduce feelings of loneliness and support the development of a positive, personal identity. They can also support young people to become more resilient to adverse situations which can help them to stay mentally healthy.
\nWhile online communities can be beneficial, they also bring associated risks. For example, members of the LGBT networks can become easy targets for abuse, discrimination, harassment and prejudice. It is therefore critical that young people understand how to keep themselves safe online and develop appropriate digital resilience to enable them to address these challenges.
\nSocial media use can allow young people to express themselves positively, letting young people put forward a positive image of themselves [8]. The problem with this is that people tend to use social media to present the best version of themselves and of their lives. This can result in others making unhealthy comparisons between their own lives and the idealised lives that are depicted on the internet, resulting in low self-esteem.
\nSocial media platforms enable young people to share creative content and express their interests and passions with others [1]. This can help to strengthen the development of a positive identity among young people and provide them with numerous opportunities to experiment with a range of interests. This is particularly important for young people who live in rural communities who may find it more challenging to develop social connections in the offline world.
\nStudents living in boarding schools benefit from using social media platforms because it enables them to maintain contact with family members and friends at home. This is particularly important because students living away from home may experience isolation and homesickness and social media platforms facilitate these connections.
\nSocial media platforms offer young people a useful tool to make, maintain or build social connections with others [1]. Additionally, research suggests that strong adolescent friendships can be enhanced by social media interactions [9]. Thus, young people can use social media to cement the friendships that they have formed in the offline world and to develop new friendships that would not have been possible in the offline word due to geographical restrictions.
\nSchools play a critical role in keeping children safe online. A well-planned digital curriculum should cover themes such as digital resilience and digital citizenship so that young people know how to respond to distressing content and how to behave responsibly online. The curriculum should also provide digital literacy skills so that children and young people have the skills to keep their own accounts safe through privacy settings, blocking perpetrators of abuse, reporting abuse and setting passwords. Schools should also support children and young people to critically engage with content they see online. They should be taught to question and interrogate content for accuracy, exploitation, abuse and discrimination.
\nSchools also play a critical role in developing young people’s mental health literacy. This should cover common mental health conditions, including stress, anxiety, depression, self-harm and cyberbullying. Educating young people about mental health is essential and reduces the stigma that has traditionally been associated with mental health conditions. Young people also need to have strategies for managing their own mental health. If their mental health is adversely affected by their experiences online, they need to be taught strategies to self-regulate their emotions and strategies to aid digital resilience. Some young people who have negative experiences online respond by closing down their social media accounts. This situates the control with the perpetrators of abuse and removes control from the victim because they are disadvantaged. Developing practical approaches to aid digital resilience in the face of adversity must be a key component of the digital curriculum that schools provide. Young people need to know how to respond to abuse, who to report it to and how to block the accounts of perpetrators. In addition, they need to be taught about the importance of maintaining secure social media accounts and how to keep themselves safe by not sharing personal information.
\nSchools need to provide a social need to provide a social media curriculum which is progressive and age appropriate. Given the prevalence of fake content online and content which has been digitally edited, young people need to be taught to critically evaluate content that appears online so that they understand the harmful effects of some content. Themes including exploitation, body-esteem and gender stereotyping can be addressed through critically evaluating online content.
\nChildren and young people often have a good understanding of the issues associated with social media because they are the users of it. Therefore, they experience the issues, sometimes frequently. Working in partnership with young people through empowering them to lead on aspects of social media education is a powerful way of developing student partnership and empowers them to be leaders. Often, young people understand the online applications better than teachers and they are acutely aware of the issues that occur online. Student-led events such as student-led workshops and conferences, which highlight the issues that relate to social media and mental health, are powerful ways of providing ownership to students. Developing digital ambassadors who act as peer mentors to younger students is also a powerful strategy for developing students’ confidence and leadership skills. Young people who need someone to talk to about the issues that they are experiencing online can be paired with a digital ambassador who can provide them with confidential advice. Processes for recruiting digital ambassadors would need to be carefully considered by schools and the scheme would need to be properly led and managed by a member of staff to monitor its effectiveness. Student-led peer mentoring schemes are valuable because some students prefer to talk to peers about the issues that they are experiencing rather than teachers or parents.
\nSchools also play a critical role in educating parents about the relationship between social media and mental health. It is important that parents understand the online applications that their children are using, and schools can play a critical role in developing their understanding. Schools can also provide guidance to parents on the signs and symptoms of mental ill health so that they are better able to identify mental health problems in their child. Schools can provide guidance to parents on how to support their child’s mental health at home and guidelines about responsible use of social media in the home. It is critical that parents understand the association between poor sleep quality, mental health and academic attainment and schools can play an important role in this. Schools also play a crucial role in developing parents’ knowledge about how to be a good social media role model for their child.
\nSchools cannot solve the problems associated with social media in isolation. This section outlines the responsibilities of parents, social media companies and advertising companies. The responsibilities of the government are also outlined.
\nParents are in a unique position to influence their child’s social media use. They should establish clear expectations about the amount of time their child spends online. However, imposing rules on children can lead to conflict and the breakdown of relationships between parents and children. It is far more effective for parents and children to negotiate the rules jointly so that young people have ownership of determining the boundaries of acceptable and unacceptable behaviour. If rules are imposed rather than negotiated it is likely that young people will find ways to break the rules and therefore adopting a top-down approach may not be the most effective way of encouraging young people to develop healthy social media use.
\nSome parents may try to restrict their child’s use of social media by installing filters or by disconnecting the internet supply at specific times of the day or week. However, young people will find ways to subvert this and policing their use of the internet in this way is unlikely to foster digital responsibility. It might be more effective for parents to talk to their child about what it means to be a digitally responsible citizen and to explain why it is important to restrict screen time, particularly during the night. Families might want to consider allocating specific time each day or week when no-one accesses technology.
\nIn addition, parents also need to be role models. They cannot expect their child to demonstrate the skills of digital citizenship and digital responsibility if they are not prepared to demonstrate these skills. It is therefore important for parents to model healthy online behaviours so that their children can then replicate these. It is also important for parents to develop their own digital literacy, so they are aware of the platforms and software that their child is interacting with. Parents also need to develop knowledge of the risks that their children are exposed to, given that these are constantly changing. If parents do not keep abreast of developments, they will not be able to support their child effectively.
\nParents should negotiate rules with their children about what constitutes appropriate use of the internet. Imposing rules on children is unlikely to be effective because young people will find ways to resist or subvert these. It is also important that parents provide their children with a degree of autonomy about their internet use. It is unlikely to be helpful if parents continually monitor what their children are doing online. However, it is reasonable for parents to set some rules for appropriate use to protect their child from harm. Examples include:
not using technology during the night;
restricting technology use during mealtimes or other social occasions;
limiting the amount of screen time which children are exposed to.
It will be more effective if young people are involved in discussions with their parents about what might constitute appropriate use of the internet.
\nSocial media companies have a responsibility to protect young people from harm. They can do this in a variety of ways by:
establishing strict and robust policies on the age at which users can access platforms;
blocking accounts of perpetrators of abuse;
reporting abuse to the police;
removing inappropriate content immediately;
filtering specific content before it goes live;
producing information to service users about responsible and safe use of social media;
generating warning messages when users have exceeded reasonable levels of screen time;
responding rapidly to reports of abuse.
This is not an exhaustive list. However, it illustrates the sorts of actions that can be adopted by social media companies to protect children and young people from harm. Companies have not responded quickly enough to reports of abuse or inappropriate content as cases of suicide in the UK suggest that social media companies have failed to protect young people from harm. The government also has a clear responsibility to hold companies to account which fail to protect children and young people from harm. Simply fining companies is not enough and will not necessarily address the problem. The government needs to take firmer action against social media companies which breach their safeguarding responsibilities.
\nIn addition, advertising companies have a responsibility to ensure that young people do not develop low body confidence. They can achieve this in a variety of ways. These include:
providing warning messages that images may have been digitally edited;
ensuring that images of bodies on products represent a range of body types, including a range of body sizes, disabled bodies and people of colour;
avoiding gender-stereotypes when advertising products;
producing warning messages about the dangers associated with product-use so that young people are aware of the risks;
portraying natural bodies without make-up on some products.
Our own research in Cambridge [10] with students in secondary schools demonstrates that they had a good understanding of the benefits and risks associated with social media. Focus groups demonstrated that the students had developed an excellent understanding of the benefits of social media and the relationship between social media use and mental ill health, including sleep deprivation, cyberbullying and low body-esteem. They had also developed a better understanding of how to keep themselves safe online. The quotes and Figure 1 below are taken from our research report [10].
\nStudents’ perspectives on social media.
\nSocial media helps you to communicate with your friends if they are far away. It makes you feel good when you get a like on your posts. (Student Y8)
\n\nYou can talk to your friends and family on social media. The disadvantages are that you can get stalked. People can create fake accounts. You can get cyber-bullied. People can hack into other people’s accounts and you might not know who is communicating with you. People can become jealous of other people’s lives and this can make you sad and depressed. (Student Y9)
\n\nSome of the pictures can be fake so people can make out that they are leading an exciting life but really, they are not, and this can make others feel worthless. (Student Y8)
\n\nSocial media results in an expectation to show the good part of your life. It can impact on others because they think you are having a good time and they might not be having such a good time. (Student Y9)
\n\nPeople make mean comments and it makes you feel bad. The bullying can be anonymous, and it reaches a larger audience. You can ignore the insults and carry on with your life. You can report the person or block them. (Student Y9)
\n\nMen are expected to be muscular. You get upset because you think “why don’t I look like that?” (Student Y8)
\n\nI realize that social media has an impact on my sleep. I find it addictive and I am always checking what friends are doing through social media and texting. (Student Y9)
\n\nI think online bullying is different to bullying in school. It is easier to say horrible things to someone through social media because you are not saying it to their face. (Student Y8)
\n\nWe can become stressed through social media because celebrities show images of being slim. This mainly affects women but now men are becoming bothered about how they look. This is stress that becomes a mental health problem. (Student Y9)
\n\nYou feel you must look as good as celebrity people because people feel you need to be as good looking otherwise you don’t get a good reputation. (Student Y8)
\n\nCyber bullying is when you post hateful messages online to directly hurt a person. (Student Y8)
\n\nSeeing slim models online (body image) can make your self-esteem feel low. (Student Y8).
\nThe students summarised the advantages and disadvantages of social media below:
\nCyberbullying is bullying which takes place in the online world, including bullying which takes place on social media. It takes multiple forms. These include:
posting hurtful comments;
posting videos which are targeted directly at a person to cause distress;
posting photographs which are designed to cause distress;
inciting others to make hurtful comments aimed at a person;
sending hurtful text messages using a mobile phone;
sending hurtful private messages to a person [11].
According to Glazzard and Mitchell [11]:
\n\nCyberbullying is fundamentally different to face-to-face bullying in several ways. Firstly, victims cannot escape from it when they are at home because it takes place on mobile phones, tablets and computers. Secondly the abuse is witnessed by a larger audience; messages are in the public domain and can be repeatedly forwarded. This can result in victims experiencing the abuse on multiple occasions, which results in further psychological distress. Thirdly, the evidence of the abuse is usually permanently stored online which means that the abuse is not erased. These messages serve as a permanent reminder of the abuse and this can result in abuse being continually experienced by the victim.
\nForms of cyberbullying are outlined below and taken from Glazzard and Mitchell [11]:
\nHarassment: Harassment is the act of sending offensive, rude, and insulting messages and being abusive. It includes nasty or degrading comments on posts, photos and in chat rooms and making offensive comments on gaming sites. Posting false and malicious things about people on the internet can be classed as harassment [11].
\nDenigration: This is when someone may send information about another person that is fake, damaging and untrue. It includes sharing photographs of someone for the purpose to ridicule and spreading fake rumours and gossip. This can be on any site online or on apps. It includes purposely altering photographs of others to ridicule and cause distress [11].
\nFlaming: Flaming is when someone purposely uses extreme and offensive language and deliberately gets into online arguments and fights. They do this to deliberately cause distress in others [11].
\nImpersonation: Impersonation is when someone hacks into someone’s email or social networking account and uses the person’s online identity to send or post vicious or embarrassing material to or about others. It also includes making up fake profiles of others [11].
\nOuting and trickery: This is when someone shares personal information about someone else or tricks someone into revealing secrets and subsequently forwards it to others. They may also do this with private images and videos too [11].
\nCyberstalking: Cyberstalking is the act of repeatedly sending messages that include threats of harm, harassment, intimidating messages, or engaging in other online activities that make a person afraid for their safety. The actions may be illegal depending on what they are doing. Cyberstalking can take place on the internet or via mobile ‘phones. Examples include:
silent calls;
insulting and threatening texts;
abusive verbal messages;
cases of stolen identities [11]
Exclusion: This is when others intentionally leave someone out of a group such as group messages, online apps, gaming sites and other online engagement. This is also a form of social bullying and is very common [11].
\nBullying by spreading rumours and gossip: Online abuse, rumours and gossip can go viral very quickly and be shared by many people within several minutes. It is not uncommon for former close friends or partners to share personal secrets about victims [11].
\nThreatening behaviour: Threatening behaviour which is directed at a victim to cause alarm and distress is a criminal offence. Taking screenshots of the evidence and reporting it is one way of challenging this [11].
\nHappy slapping: This is an incident where a person is assaulted while other people take photographs or videos on their mobile phones. The pictures or videos are then circulated by mobile phone or uploaded on the internet [11].
\nGrooming: Grooming is when someone builds an emotional connection with a child to gain their trust for the purposes of abuse and exploitation. It is conducted by strangers (or new “friends”) and may include:
pressurising someone to do something they do not wish to do;
making someone take their clothes off;
pressurising someone to engage in sexual conversations;
pressurising someone to take naked photographs of themselves;
making someone engage in sexual activity via the internet [11].
Groomers may spend a long time establishing a “relationship” with the victim by using the following strategies:
pretending to be someone they are not, for example, saying they are the same age online;
offering advice or understanding;
buying gifts;
giving the child attention;
using their professional position or reputation;
giving compliments;
taking them on trips, outings or holidays [11].
Inappropriate images: It is very easy to save any pictures of anyone on any site and upload them to the internet. Uploading pictures of someone to cause distress is a form of cyberbullying. This also includes digitally altering pictures to embarrass someone [11].
\nBystander effect: Witnessing cyberbullying and doing nothing about it is not acceptable. Some people are worried about getting involved but victims of bullying need brave witnesses to make a stand. Perpetrators of bullying thrive when they have an audience. Making a stand against what they are doing is an important way to reduce their power. Most sites now operate a reporting facility so that online abuse can be reported and addressed. Bystanders are not innocent. They have a responsibility to report abuse that they witness [11].
\nThe following text is taken from our blog [12].
\n\nResearch from Queensland University of Technology has identified that half of young people aged 18–24 are less productive and more tired because of their mobile phones. Scientists have adopted the term “technoference” to describe the way that mobile phones intrude on and interrupt everyday conversations and the way they interrupt other aspects of people’s daily lives.
\n\nIt is worrying that family life is being interrupted by technology. While technology has significant benefits, continual use of technology can impact detrimentally on the quality of people’s interactions and conversations. We live in a society where people are constantly attached to their technology. People interact with technology on public transport, in meetings and during leisure time rather than engaging in productive, meaningful conversations. It seems that people would rather interact with a phone rather than having a conversation and while this is not necessarily a problem in some contexts, it can have a negative impact in other contexts. For example, young children require social interaction with adults. This allows them to develop secure attachments with significant others, it enables them to learn about the world and through conversation children are exposed to language. Exposure to language underpins reading and writing development. Children who have rich exposure to language become better readers, better writers and understand far better what they are reading. Lack of exposure to language can impact detrimentally on the structure of the brain. This can create reading difficulties and even lead to difficulties which are consistent with dyslexia, even though the difficulties may not have a genetic origin. The brain is malleable. It is responsive to environmental influences and lack of exposure to language can impact on phonological and phonemic awareness. Both of these skills play a critical role in reading development. Interacting with technology can restrict opportunities for communication between babies, children and their parents and can interrupt the flow of normal conversation.\n
\n\nIt would appear that adolescents seem to be attached to their phones during the night. They are desperate to network and keep up-to-date with their online peers. This results in broken sleep and tiredness during the school day. Adolescents need approximately 8–10 hours sleep but our research demonstrates that some get as little as 2 hours sleep. These students attend school in a state of exhaustion. They are too tired to concentrate and it affects their learning and their behaviour. Disengagement in lessons results in them falling behind in their schoolwork and they then develop other problems such as low confidence and low self-worth.
\n\nReal-time social connections are vital for positive wellbeing. Schools play a key role in teaching young people about how to stay healthy and in particular, the need for sleep. However, parents also play a critical role in supporting young people to develop positive habits through setting boundaries. Examples of boundaries might include restricting access to technology in bedrooms and at mealtimes. Also, parents need to be good role models by ensuring that they do not allow technology to interrupt conversations and other daily experiences.\n
\nStatistics demonstrate the risks of internet use on young people’s lives. Key statistics are summarised below [13]:
year on year increases in the numbers and rates of police-recorded online child sexual offences in England and Wales and Northern Ireland
increases in police-recorded offences of obscene publications or indecent photos in all four UK nations over the last 5 years
increases in the number of URLs confirmed by the Internet Watch Foundation (IWF) as containing child sexual abuse imagery since 2015
less than half of children aged 12–15 say they know how to change their settings to control who can view their social media
the majority of parents, carers and members of the public agree that social networks should have a legal responsibility to keep children safe on their platforms.
Additionally:
a total of 5161 crimes of sexual communication with a child have been recorded in 18 months [14];
in 2019 there has been almost a 50% increase in offence in offences recorded in latest 6 months compared to same period in previous year [14];
in 2010 there has been a 200% rise in recorded instances in the use of Instagram to target and abuse children over the same time period [14];
there have been over 5000 online grooming offences recorded in 18 months [14].
Social media use can have a detrimental impact on children and young people’s mental health. It can result in anxiety, depression, body image concerns, self-harm, substance abuse and even death. However, for young people social media is a tool for networking, keeping in touch with friends, exchanging information, a source of support and advice and a rich source of knowledge. Preventing children and young people from using social media is not an appropriate solution, given all the benefits that come with it. Schools, parents and the digital industry need to do all they can to keep children safe from harm through adopting a proactive approach rather than a reactive approach when crises occur.
\nWe wish to thank Leeds Beckett University and the Carnegie Centre of Excellence for Mental Health in Schools for facilitating this research.
\nThe authors declare no conflict of interest.
"I work with IntechOpen for a number of reasons: their professionalism, their mission in support of Open Access publishing, and the quality of their peer-reviewed publications, but also because they believe in equality. Throughout the world, we are seeing progress in attracting, retaining, and promoting women in STEMM. IntechOpen are certainly supporting this work globally by empowering all scientists and ensuring that women are encouraged and enabled to publish and take leading roles within the scientific community." Dr. Catrin Rutland, University of Nottingham, UK
",metaTitle:"Advantages of Publishing with IntechOpen",metaDescription:"We have more than a decade of experience in Open Access publishing. \n\n ",metaKeywords:null,canonicalURL:null,contentRaw:'[{"type":"htmlEditorComponent","content":"We have more than a decade of experience in Open Access publishing. The advantages of publishing with IntechOpen include:
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\\n\\nOur expertise – We’ve published more than 4,500 books by more than 118,000 authors and editors.
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\\n\\nOur services – The support we offer our authors and editors is second to none. Each book in our program receives the following:
\\n\\nOur end-to-end publishing service frees our authors and editors to focus on what matters: research. We empower them to shape their fields and connect with the global scientific community.
\\n\\n"In developing countries until now, advancement in science has been very limited, because insufficient economic resources are dedicated to science and education. These limitations are more marked when the scientists are women. In order to develop science in the poorest countries and decrease the gender gap that exists in scientific fields, Open Access networks like IntechOpen are essential. Free access to scientific research could contribute to ameliorating difficult life conditions and breaking down barriers." Marquidia Pacheco, National Institute for Nuclear Research (ININ), Mexico
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\n\nOur platform – IntechOpen is the world’s leading publisher of OA books, built by scientists, for scientists.
\n\nOur reputation – Everything we publish goes through a two-stage peer review process. We’re proud to count Nobel laureates among our esteemed authors. We meet European Commission standards for funding, and the research we’ve published has been funded by the Bill and Melinda Gates Foundation and the Wellcome Trust, among others. IntechOpen is a member of all relevant trade associations (including the STM Association and the Association of Learned and Professional Society Publishers) and has a selection of books indexed in Web of Science's Book Citation Index.
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\n\nOur end-to-end publishing service frees our authors and editors to focus on what matters: research. We empower them to shape their fields and connect with the global scientific community.
\n\n"In developing countries until now, advancement in science has been very limited, because insufficient economic resources are dedicated to science and education. These limitations are more marked when the scientists are women. In order to develop science in the poorest countries and decrease the gender gap that exists in scientific fields, Open Access networks like IntechOpen are essential. Free access to scientific research could contribute to ameliorating difficult life conditions and breaking down barriers." Marquidia Pacheco, National Institute for Nuclear Research (ININ), Mexico
\n\nInterested? Contact Ana Pantar (book.idea@intechopen.com) for more information.
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She performed (inter)national tasks as vice-president of the Concilium Anaesthesia and related committees. \nShe performed research in several fields, with over 100 publications in (inter)national journals and numerous papers on scientific conferences. \nShe received several awards and is a member of Honour of the Dutch Society of Anaesthesia.",institutionString:null,institution:{name:"Albert Schweitzer Hospital",country:{name:"Gabon"}}},{id:"83089",title:"Prof.",name:"Aaron",middleName:null,surname:"Ojule",slug:"aaron-ojule",fullName:"Aaron Ojule",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Port Harcourt",country:{name:"Nigeria"}}},{id:"295748",title:"Mr.",name:"Abayomi",middleName:null,surname:"Modupe",slug:"abayomi-modupe",fullName:"Abayomi Modupe",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/no_image.jpg",biography:null,institutionString:null,institution:{name:"Landmark University",country:{name:"Nigeria"}}},{id:"94191",title:"Prof.",name:"Abbas",middleName:null,surname:"Moustafa",slug:"abbas-moustafa",fullName:"Abbas Moustafa",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/94191/images/96_n.jpg",biography:"Prof. Moustafa got his doctoral degree in earthquake engineering and structural safety from Indian Institute of Science in 2002. He is currently an associate professor at Department of Civil Engineering, Minia University, Egypt and the chairman of Department of Civil Engineering, High Institute of Engineering and Technology, Giza, Egypt. He is also a consultant engineer and head of structural group at Hamza Associates, Giza, Egypt. Dr. Moustafa was a senior research associate at Vanderbilt University and a JSPS fellow at Kyoto and Nagasaki Universities. He has more than 40 research papers published in international journals and conferences. He acts as an editorial board member and a reviewer for several regional and international journals. His research interest includes earthquake engineering, seismic design, nonlinear dynamics, random vibration, structural reliability, structural health monitoring and uncertainty modeling.",institutionString:null,institution:{name:"Minia University",country:{name:"Egypt"}}},{id:"84562",title:"Dr.",name:"Abbyssinia",middleName:null,surname:"Mushunje",slug:"abbyssinia-mushunje",fullName:"Abbyssinia Mushunje",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"University of Fort Hare",country:{name:"South Africa"}}},{id:"202206",title:"Associate Prof.",name:"Abd Elmoniem",middleName:"Ahmed",surname:"Elzain",slug:"abd-elmoniem-elzain",fullName:"Abd Elmoniem Elzain",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Kassala University",country:{name:"Sudan"}}},{id:"98127",title:"Dr.",name:"Abdallah",middleName:null,surname:"Handoura",slug:"abdallah-handoura",fullName:"Abdallah Handoura",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"École Supérieure des Télécommunications",country:{name:"Morocco"}}},{id:"91404",title:"Prof.",name:"Abdecharif",middleName:null,surname:"Boumaza",slug:"abdecharif-boumaza",fullName:"Abdecharif Boumaza",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Abbès Laghrour University of Khenchela",country:{name:"Algeria"}}},{id:"105795",title:"Prof.",name:"Abdel Ghani",middleName:null,surname:"Aissaoui",slug:"abdel-ghani-aissaoui",fullName:"Abdel Ghani Aissaoui",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/105795/images/system/105795.jpeg",biography:"Abdel Ghani AISSAOUI is a Full Professor of electrical engineering at University of Bechar (ALGERIA). He was born in 1969 in Naama, Algeria. He received his BS degree in 1993, the MS degree in 1997, the PhD degree in 2007 from the Electrical Engineering Institute of Djilali Liabes University of Sidi Bel Abbes (ALGERIA). He is an active member of IRECOM (Interaction Réseaux Electriques - COnvertisseurs Machines) Laboratory and IEEE senior member. He is an editor member for many international journals (IJET, RSE, MER, IJECE, etc.), he serves as a reviewer in international journals (IJAC, ECPS, COMPEL, etc.). He serves as member in technical committee (TPC) and reviewer in international conferences (CHUSER 2011, SHUSER 2012, PECON 2012, SAI 2013, SCSE2013, SDM2014, SEB2014, PEMC2014, PEAM2014, SEB (2014, 2015), ICRERA (2015, 2016, 2017, 2018,-2019), etc.). His current research interest includes power electronics, control of electrical machines, artificial intelligence and Renewable energies.",institutionString:"University of Béchar",institution:{name:"University of Béchar",country:{name:"Algeria"}}},{id:"99749",title:"Dr.",name:"Abdel Hafid",middleName:null,surname:"Essadki",slug:"abdel-hafid-essadki",fullName:"Abdel Hafid Essadki",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"École Nationale Supérieure de Technologie",country:{name:"Algeria"}}},{id:"101208",title:"Prof.",name:"Abdel Karim",middleName:"Mohamad",surname:"El Hemaly",slug:"abdel-karim-el-hemaly",fullName:"Abdel Karim El Hemaly",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/101208/images/733_n.jpg",biography:"OBGYN.net Editorial Advisor Urogynecology.\nAbdel Karim M. A. El-Hemaly, MRCOG, FRCS � Egypt.\n \nAbdel Karim M. A. El-Hemaly\nProfessor OB/GYN & Urogynecology\nFaculty of medicine, Al-Azhar University \nPersonal Information: \nMarried with two children\nWife: Professor Laila A. Moussa MD.\nSons: Mohamad A. M. El-Hemaly Jr. MD. Died March 25-2007\nMostafa A. M. El-Hemaly, Computer Scientist working at Microsoft Seatle, USA. \nQualifications: \n1.\tM.B.-Bch Cairo Univ. June 1963. \n2.\tDiploma Ob./Gyn. Cairo Univ. April 1966. \n3.\tDiploma Surgery Cairo Univ. Oct. 1966. \n4.\tMRCOG London Feb. 1975. \n5.\tF.R.C.S. Glasgow June 1976. \n6.\tPopulation Study Johns Hopkins 1981. \n7.\tGyn. Oncology Johns Hopkins 1983. \n8.\tAdvanced Laparoscopic Surgery, with Prof. Paulson, Alexandria, Virginia USA 1993. \nSocieties & Associations: \n1.\t Member of the Royal College of Ob./Gyn. London. \n2.\tFellow of the Royal College of Surgeons Glasgow UK. \n3.\tMember of the advisory board on urogyn. FIGO. \n4.\tMember of the New York Academy of Sciences. \n5.\tMember of the American Association for the Advancement of Science. \n6.\tFeatured in �Who is Who in the World� from the 16th edition to the 20th edition. \n7.\tFeatured in �Who is Who in Science and Engineering� in the 7th edition. \n8.\tMember of the Egyptian Fertility & Sterility Society. \n9.\tMember of the Egyptian Society of Ob./Gyn. \n10.\tMember of the Egyptian Society of Urogyn. \n\nScientific Publications & Communications:\n1- Abdel Karim M. El Hemaly*, Ibrahim M. Kandil, Asim Kurjak, Ahmad G. Serour, Laila A. S. Mousa, Amr M. Zaied, Khalid Z. El Sheikha. \nImaging the Internal Urethral Sphincter and the Vagina in Normal Women and Women Suffering from Stress Urinary Incontinence and Vaginal Prolapse. Gynaecologia Et Perinatologia, Vol18, No 4; 169-286 October-December 2009.\n2- Abdel Karim M. El Hemaly*, Laila A. S. Mousa Ibrahim M. Kandil, Fatma S. El Sokkary, Ahmad G. Serour, Hossam Hussein.\nFecal Incontinence, A Novel Concept: The Role of the internal Anal sphincter (IAS) in defecation and fecal incontinence. Gynaecologia Et Perinatologia, Vol19, No 2; 79-85 April -June 2010.\n3- Abdel Karim M. El Hemaly*, Laila A. S. Mousa Ibrahim M. Kandil, Fatma S. El Sokkary, Ahmad G. Serour, Hossam Hussein.\nSurgical Treatment of Stress Urinary Incontinence, Fecal Incontinence and Vaginal Prolapse By A Novel Operation \n"Urethro-Ano-Vaginoplasty"\n Gynaecologia Et Perinatologia, Vol19, No 3; 129-188 July-September 2010.\n4- Abdel Karim M. El Hemaly*, Ibrahim M. Kandil, Laila A. S. Mousa and Mohamad A.K.M.El Hemaly.\nUrethro-vaginoplasty, an innovated operation for the treatment of: Stress Urinary Incontinence (SUI), Detursor Overactivity (DO), Mixed Urinary Incontinence and Anterior Vaginal Wall Descent. \nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/ urethro-vaginoplasty_01\n\n5- Abdel Karim M. El Hemaly, Ibrahim M Kandil, Mohamed M. Radwan.\n Urethro-raphy a new technique for surgical management of Stress Urinary Incontinence.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/\nnew-tech-urethro\n\n6- Abdel Karim M. El Hemaly, Ibrahim M Kandil, Mohamad A. Rizk, Nabil Abdel Maksoud H., Mohamad M. Radwan, Khalid Z. El Shieka, Mohamad A. K. M. El Hemaly, and Ahmad T. El Saban.\nUrethro-raphy The New Operation for the treatment of stress urinary incontinence, SUI, detrusor instability, DI, and mixed-type of urinary incontinence; short and long term results. \nhttp://www.obgyn.net/urogyn/urogyn.asp?page=urogyn/articles/\nurethroraphy-09280\n\n7-Abdel Karim M. El Hemaly, Ibrahim M Kandil, and Bahaa E. El Mohamady. Menopause, and Voiding troubles. \nhttp://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly03/el-hemaly03-ss\n\n8-El Hemaly AKMA, Mousa L.A. Micturition and Urinary\tContinence. Int J Gynecol Obstet 1996; 42: 291-2. \n\n9-Abdel Karim M. El Hemaly.\n Urinary incontinence in gynecology, a review article.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/abs-urinary_incotinence_gyn_ehemaly \n\n10-El Hemaly AKMA. Nocturnal Enuresis: Pathogenesis and Treatment. \nInt Urogynecol J Pelvic Floor Dysfunct 1998;9: 129-31.\n \n11-El Hemaly AKMA, Mousa L.A.E. Stress Urinary Incontinence, a New Concept. Eur J Obstet Gynecol Reprod Biol 1996; 68: 129-35. \n\n12- El Hemaly AKMA, Kandil I. M. Stress Urinary Incontinence SUI facts and fiction. Is SUI a puzzle?! http://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly/el-hemaly-ss\n\n13-Abdel Karim El Hemaly, Nabil Abdel Maksoud, Laila A. Mousa, Ibrahim M. Kandil, Asem Anwar, M.A.K El Hemaly and Bahaa E. El Mohamady. \nEvidence based Facts on the Pathogenesis and Management of SUI. http://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly02/el-hemaly02-ss\n\n14- Abdel Karim M. El Hemaly*, Ibrahim M. Kandil, Mohamad A. Rizk and Mohamad A.K.M.El Hemaly.\n Urethro-plasty, a Novel Operation based on a New Concept, for the Treatment of Stress Urinary Incontinence, S.U.I., Detrusor Instability, D.I., and Mixed-type of Urinary Incontinence.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/urethro-plasty_01\n\n15-Ibrahim M. Kandil, Abdel Karim M. El Hemaly, Mohamad M. Radwan: Ultrasonic Assessment of the Internal Urethral Sphincter in Stress Urinary Incontinence. The Internet Journal of Gynecology and Obstetrics. 2003. Volume 2 Number 1. \n\n\n16-Abdel Karim M. El Hemaly. Nocturnal Enureses: A Novel Concept on its pathogenesis and Treatment.\nhttp://www.obgyn.net/urogynecolgy/?page=articles/nocturnal_enuresis\n\n17- Abdel Karim M. El Hemaly. Nocturnal Enureses: An Update on the pathogenesis and Treatment.\nhttp://www.obgyn.net/urogynecology/?page=/ENHLIDH/PUBD/FEATURES/\nPresentations/ Nocturnal_Enuresis/nocturnal_enuresis\n\n18-Maternal Mortality in Egypt, a cry for help and attention. The Second International Conference of the African Society of Organization & Gestosis, 1998, 3rd Annual International Conference of Ob/Gyn Department � Sohag Faculty of Medicine University. Feb. 11-13. Luxor, Egypt. \n19-Postmenopausal Osteprosis. The 2nd annual conference of Health Insurance Organization on Family Planning and its role in primary health care. Zagaziz, Egypt, February 26-27, 1997, Center of Complementary Services for Maternity and childhood care. \n20-Laparoscopic Assisted vaginal hysterectomy. 10th International Annual Congress Modern Trends in Reproductive Techniques 23-24 March 1995. Alexandria, Egypt. \n21-Immunological Studies in Pre-eclamptic Toxaemia. Proceedings of 10th Annual Ain Shams Medical Congress. Cairo, Egypt, March 6-10, 1987. \n22-Socio-demographic factorse affecting acceptability of the long-acting contraceptive injections in a rural Egyptian community. Journal of Biosocial Science 29:305, 1987. \n23-Plasma fibronectin levels hypertension during pregnancy. The Journal of the Egypt. Soc. of Ob./Gyn. 13:1, 17-21, Jan. 1987. \n24-Effect of smoking on pregnancy. Journal of Egypt. Soc. of Ob./Gyn. 12:3, 111-121, Sept 1986. \n25-Socio-demographic aspects of nausea and vomiting in early pregnancy. Journal of the Egypt. Soc. of Ob./Gyn. 12:3, 35-42, Sept. 1986. \n26-Effect of intrapartum oxygen inhalation on maternofetal blood gases and pH. Journal of the Egypt. Soc. of Ob./Gyn. 12:3, 57-64, Sept. 1986. \n27-The effect of severe pre-eclampsia on serum transaminases. The Egypt. J. Med. Sci. 7(2): 479-485, 1986. \n28-A study of placental immunoreceptors in pre-eclampsia. The Egypt. J. Med. Sci. 7(2): 211-216, 1986. \n29-Serum human placental lactogen (hpl) in normal, toxaemic and diabetic pregnant women, during pregnancy and its relation to the outcome of pregnancy. Journal of the Egypt. Soc. of Ob./Gyn. 12:2, 11-23, May 1986. \n30-Pregnancy specific B1 Glycoprotein and free estriol in the serum of normal, toxaemic and diabetic pregnant women during pregnancy and after delivery. Journal of the Egypt. Soc. of Ob./Gyn. 12:1, 63-70, Jan. 1986. Also was accepted and presented at Xith World Congress of Gynecology and Obstetrics, Berlin (West), September 15-20, 1985. \n31-Pregnancy and labor in women over the age of forty years. Accepted and presented at Al-Azhar International Medical Conference, Cairo 28-31 Dec. 1985. \n32-Effect of Copper T intra-uterine device on cervico-vaginal flora. Int. J. Gynaecol. Obstet. 23:2, 153-156, April 1985. \n33-Factors affecting the occurrence of post-Caesarean section febrile morbidity. Population Sciences, 6, 139-149, 1985. \n34-Pre-eclamptic toxaemia and its relation to H.L.A. system. Population Sciences, 6, 131-139, 1985. \n35-The menstrual pattern and occurrence of pregnancy one year after discontinuation of Depo-medroxy progesterone acetate as a postpartum contraceptive. Population Sciences, 6, 105-111, 1985. \n36-The menstrual pattern and side effects of Depo-medroxy progesterone acetate as postpartum contraceptive. Population Sciences, 6, 97-105, 1985. \n37-Actinomyces in the vaginas of women with and without intrauterine contraceptive devices. Population Sciences, 6, 77-85, 1985. \n38-Comparative efficacy of ibuprofen and etamsylate in the treatment of I.U.D. menorrhagia. Population Sciences, 6, 63-77, 1985. \n39-Changes in cervical mucus copper and zinc in women using I.U.D.�s. Population Sciences, 6, 35-41, 1985. \n40-Histochemical study of the endometrium of infertile women. Egypt. J. Histol. 8(1) 63-66, 1985. \n41-Genital flora in pre- and post-menopausal women. Egypt. J. Med. Sci. 4(2), 165-172, 1983. \n42-Evaluation of the vaginal rugae and thickness in 8 different groups. Journal of the Egypt. Soc. of Ob./Gyn. 9:2, 101-114, May 1983. \n43-The effect of menopausal status and conjugated oestrogen therapy on serum cholesterol, triglycerides and electrophoretic lipoprotein patterns. Al-Azhar Medical Journal, 12:2, 113-119, April 1983. \n44-Laparoscopic ventrosuspension: A New Technique. Int. J. Gynaecol. Obstet., 20, 129-31, 1982. \n45-The laparoscope: A useful diagnostic tool in general surgery. Al-Azhar Medical Journal, 11:4, 397-401, Oct. 1982. \n46-The value of the laparoscope in the diagnosis of polycystic ovary. Al-Azhar Medical Journal, 11:2, 153-159, April 1982. \n47-An anaesthetic approach to the management of eclampsia. Ain Shams Medical Journal, accepted for publication 1981. \n48-Laparoscopy on patients with previous lower abdominal surgery. Fertility management edited by E. Osman and M. Wahba 1981. \n49-Heart diseases with pregnancy. Population Sciences, 11, 121-130, 1981. \n50-A study of the biosocial factors affecting perinatal mortality in an Egyptian maternity hospital. Population Sciences, 6, 71-90, 1981. \n51-Pregnancy Wastage. Journal of the Egypt. Soc. of Ob./Gyn. 11:3, 57-67, Sept. 1980. \n52-Analysis of maternal deaths in Egyptian maternity hospitals. Population Sciences, 1, 59-65, 1979. \nArticles published on OBGYN.net: \n1- Abdel Karim M. El Hemaly*, Ibrahim M. Kandil, Laila A. S. Mousa and Mohamad A.K.M.El Hemaly.\nUrethro-vaginoplasty, an innovated operation for the treatment of: Stress Urinary Incontinence (SUI), Detursor Overactivity (DO), Mixed Urinary Incontinence and Anterior Vaginal Wall Descent. \nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/ urethro-vaginoplasty_01\n\n2- Abdel Karim M. El Hemaly, Ibrahim M Kandil, Mohamed M. Radwan.\n Urethro-raphy a new technique for surgical management of Stress Urinary Incontinence.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/\nnew-tech-urethro\n\n3- Abdel Karim M. El Hemaly, Ibrahim M Kandil, Mohamad A. Rizk, Nabil Abdel Maksoud H., Mohamad M. Radwan, Khalid Z. El Shieka, Mohamad A. K. M. El Hemaly, and Ahmad T. El Saban.\nUrethro-raphy The New Operation for the treatment of stress urinary incontinence, SUI, detrusor instability, DI, and mixed-type of urinary incontinence; short and long term results. \nhttp://www.obgyn.net/urogyn/urogyn.asp?page=urogyn/articles/\nurethroraphy-09280\n\n4-Abdel Karim M. El Hemaly, Ibrahim M Kandil, and Bahaa E. El Mohamady. Menopause, and Voiding troubles. \nhttp://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly03/el-hemaly03-ss\n\n5-El Hemaly AKMA, Mousa L.A. Micturition and Urinary\tContinence. Int J Gynecol Obstet 1996; 42: 291-2. \n\n6-Abdel Karim M. El Hemaly.\n Urinary incontinence in gynecology, a review article.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/abs-urinary_incotinence_gyn_ehemaly \n\n7-El Hemaly AKMA. Nocturnal Enuresis: Pathogenesis and Treatment. \nInt Urogynecol J Pelvic Floor Dysfunct 1998;9: 129-31.\n \n8-El Hemaly AKMA, Mousa L.A.E. Stress Urinary Incontinence, a New Concept. Eur J Obstet Gynecol Reprod Biol 1996; 68: 129-35. \n\n9- El Hemaly AKMA, Kandil I. M. Stress Urinary Incontinence SUI facts and fiction. Is SUI a puzzle?! http://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly/el-hemaly-ss\n\n10-Abdel Karim El Hemaly, Nabil Abdel Maksoud, Laila A. Mousa, Ibrahim M. Kandil, Asem Anwar, M.A.K El Hemaly and Bahaa E. El Mohamady. \nEvidence based Facts on the Pathogenesis and Management of SUI. http://www.obgyn.net/displayppt.asp?page=/English/pubs/features/presentations/El-Hemaly02/el-hemaly02-ss\n\n11- Abdel Karim M. El Hemaly*, Ibrahim M. Kandil, Mohamad A. Rizk and Mohamad A.K.M.El Hemaly.\n Urethro-plasty, a Novel Operation based on a New Concept, for the Treatment of Stress Urinary Incontinence, S.U.I., Detrusor Instability, D.I., and Mixed-type of Urinary Incontinence.\nhttp://www.obgyn.net/urogyn/urogyn.asp?page=/urogyn/articles/urethro-plasty_01\n\n12-Ibrahim M. Kandil, Abdel Karim M. El Hemaly, Mohamad M. Radwan: Ultrasonic Assessment of the Internal Urethral Sphincter in Stress Urinary Incontinence. The Internet Journal of Gynecology and Obstetrics. 2003. Volume 2 Number 1. \n\n13-Abdel Karim M. El Hemaly. Nocturnal Enureses: A Novel Concept on its pathogenesis and Treatment.\nhttp://www.obgyn.net/urogynecolgy/?page=articles/nocturnal_enuresis\n\n14- Abdel Karim M. El Hemaly. 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