Volatile compounds emitted by fully open flower of
Orchidaceae is the largest family of angiosperms with an estimation of 17000 to 35000 species in 880 genera (Chai & Yu, 2007). In Malaysia, more than 230 orchid genera and 4000 species had been discovered (Go et al., 2012). In Penisular Malaysia, a total of 898 species in 143 genera are currently recognised (Go et al., 2010). The amazing vast diversity of types and forms enable the Orchidaceae to be successfully distributed and colonised almost every habitats worldwide (Arditti, 1992). As a result of selective forces from evolution, orchids are found to be evolved from its ancestral forms and adapted well to their present habitats (Aceto & Gaudio, 2011). Associated with their diverse floral morphology and physiology properties, they have drawn the attention of botanists and scientists for centuries. There are orchids which resemble moths (
Similar to other angiosperms, two whorls of perianth segment can also be found in orchids. The outer and inner whorls of the orchid flowers consist of three petals and three sepals. The labellum or lip (one of the petals), is distinctly evolved from the other two morphologically and physically (Arditti, 1992). The lifespan of opened-orchid flowers can range from as short as one day to as long as 270 days (Micheneau et al., 2008).
Most orchids are epiphytic that obtain their support from trees but not for nutrition while the rest are terrestrial plants (Rada & Jaimez, 1992). Orchids’ cultivation was famous since 5000 years ago in China where
Orchid hybrids cultivation started since 1856 by John Dominy (http://www/ionopsis.com/hybridization.htm).
The export of orchids from Malaysia, Thailand and Singapore contributed RM200 million annually in the world floral (Ooi, 2005). According to the Japan Florists’ Telegraph Delivery Association, cut flower in Japan constituted around 13.3% of the imported market in the year 2010 with Malaysia having the largest market share of imports, which accounted for 23.4% (7,648 million yen), followed by Columbia at 19.2% and China at 10.4%. Malaysian orchids consisted of 8.1% of the imported cut flower orchid to Japan.
1.2. Vandaceous orchids
Tropical Asia is the native home for approximately 50 vandaceous orchids species. They are distributed in Sri Lanka and southern India to New Guinea, northern Australia and Solomon Islands, and north to China, Taiwan and the Philippines. Thailand was found to be predominated with 11 vanda species. In Thailand, vanda is a vital commercial orchid. Most of them exhibit monopodial growth where their leaves are varies according to habitat. Vandas have many different colours, and majority are yellowish-brown with dark brown spots.
The vanda has been designated as the ‘Queen Orchid of the East’ due to its robust and large rounded flowers (Teo, 1981). Most of the vandaceous orchids prefer sunlight but some are well adapted to shady areas. Like any other tropical orchids, they require warm temperature with good aeration. The vandaceous orchids take around three and half to 10 years to become mature flowering plants (Kishor et al., 2008). Once matured, this orchid genus blooms every two to three months with the flowers lasting two to three weeks. As a result of land development, 28 orchid species have been listed as endangered species on Appendix II of the Convention on International Trade in Endangered Species (CITES) and prohibited from worldwide export. Among those orchids, two belong to vandas (
It is impossible to differentiate or identify an orchid species based on the vegetative parts of the plant alone. Hence, a convenient and flower-independent method to allow quick assessment of a given orchid vegetative specimen for species confirmation can be achieved with the help of molecular markers. To date, approximately 50 species are registered under vandas in the Royal Horticultural Society (RHS) database due to their commercial importance.
Vanda Mimi Palmer
Orchids cultivation entails hard work as the orchids can be easily infected by orchid-infecting viruses. More than 50 orchid-infecting viruses have been detected worldwide, with cymbidium mosaic virus (CymMV) and odontoglossum ringspot tobamovirus (ORSV) infections being the most prevalent (Sherpa et al., 2007). Infected orchid cultivars usually exhibit blossoms with brown necrotic streaks and other necrosis symptoms. Infected flowers are smaller in size and poorer in quality. This has caused severe economic damages in the cut flower and potted plant industry (Ajjikuttira et al., 2002; Sherpa et al., 2007; Vejaratpimol et al., 1999). CymMV infection is dominant and extremely stable in Orchidaceae, and it was found to be prevalent in VMP. A screen of our VMP cDNA library revealed a 1.6% contamination rate with CymMV genes (Teh et al., 2011). Markers might be useful in facilitating the screening of virus-infected stock plants to minimise losses incurred in the floral industry. So far, the identification of CymMV infection is done through serological, bioassay or electron microscopy. Those techniques include enzyme-linked immunosorbent assays (ELISA), dot-blot immunoassay (DBIA), rapid immunofilter paper assay (RIPA), immunosorbent electron microscope (ISEM), DIG-labelled cRNA probes, reverse transcription polymerase chain reaction (RT-PCR), quartz crystal microbalance-based DNA biosensors and TaqMan real-time quantitative RT-PCR (Eun et al., 2002; Eun & Wong,2000; Hsu et al., 1992; Hu
2. Importance of floral fragrance
Floral fragrance plays various functions in both the floral and vegetative organs. Fragrance is defined as a highly complex component of floral phenotype for its dynamic patterns of emission and chemical composition (Raguso, 2008). Due to their restriction to specific lineages and interactions in species-specific ecology, these have led to their designation as specialized or secondary metabolites (Pichersky et al., 2006).
Floral fragrance has a significant impact in plant reproduction as it is a selective attractant in a variety of animal pollinators especially insects. Pollinators such as bees, butterflies and moths are able to discriminate visitation on plants based on the compositions of the floral scent. This plant-insect interaction has led to many successful pollinations and development of fruits in many crop species (Majetic et al., 2009; Shuttleworth & Johnson, 2009).
Anti-microbial or anti-herbivore properties of floral volatiles could be used by plants to protect their vital floral reproductive parts from potential predators. Two types of plant defences can be characterised based on floral volatiles property, that are direct and indirect defences such as herbivore-induced volatiles signals, and visual and olfactory floral signals to attract pollinators (Schiestl, 2010). Indirect plant defences protect the plants by minimizing damage to plant tissues through attracting arthropods that prey or parasitize the herbivores. This general property has been reported in cabbage (Park et al., 2005),
Through the discovery of pollinator-attracting floral scents as the source of olfactory pleasure since ancient times, humans had figured out unique values in certain types of floral scents and exploited them to cultivate and propagate specific plant species. For centuries, flowers with vibrant colours and scents have been used by people to enhance their beauty and this was seen in almost all major civilizations. Large numbers of aromatic plants have been used as flavourings, preservatives and herbal remedies (Pichersky et al., 2006). Their economic importance also relies on their petals which are found to be the main site of natural fragrances and flavourings in most of the plants (Baudino et al., 2007).
2.1. Fragrance biosynthesis pathways
Plants are known with the capability of synthesizing many volatile metabolites, either primary or secondary metabolites with variety of functions (Pichersky et al., 2006). However, volatile esters formation is not restricted to plant kingdom but also in yeast and fungi especially in the fermentation industry (Beekwilder et al., 2004). Floral scent is made up of a complex mixture of low-molecular-weight lipophilic compounds which are typically liquids with high vapour pressures (Vainstein et al., 2001). With the discovery of novel techniques including gas chromatography-nuclear magnetic resonance (GC-NMR), gas chromatography-mass spectrometry (GC-MS), headspace based techniques in volatiles detection and analyses, the number of identified volatile compounds has increased tremendously (Gonzalez-Mas et al., 2008; Mohd-Hairul et al., 2010; Nojima et al., 2011).
Most of the volatile compounds are derived from three major biosynthesis pathways: phenylpropanoids, fatty acid derivatives and terpenoids. They are thus classified into three major categories: terpenes, lipid derivatives and aromatic compounds. The terpenes are the largest class of plant volatiles, which consist of monoterpene alcohols and sesquiterpenes. There are also other terpene derivatives like ketones that are present in low quantities but have significant contributions to the floral fragrance. The second largest family of plant volatiles is the aromatic compounds. Most of them are derived from the intermediates in the benzenoid biosynthesis pathway that resulted in the synthesis of phenylalanine from the shikimate pathway, followed by a wide range of primary metabolites (eugenol, a lignin precursor, is one of them) and secondary non-volatile compounds (this was well reviewed in Bick & Lange, 2003; Pichersky & Dudareva, 2007).
The third category of plant volatiles is the lipid derivatives from the oxidative cleavage and decarboxylation of a variety of fatty acids which lead to shorter-chain volatiles with aldehyde and ketone moieties formation (reviewed in Baysal & Demirdoven, 2007). There are also other plant volatiles especially those with nitrogen or sulfur, which are produced through the cleavage of modified amino acids or their precursors (Cherri et al., 2007; Pichersky et al., 2006).
2.2. Discovery of fragrance-related genes
Due to the invisibility of floral scents and its dynamic nature, the study on flower scent is limited. There is no convenient plant model system that allows chemical and biochemical studies on floral scents. The well-established
‘Scent’ enzymes can be identified through
For more than 400 orchids (including both species and hybrids) that were discovered to emit fragrance (Frowine, 2005), in-depth scientific studies on orchid fragrance barely covered 2 percents of the fragrant orchids. Sadly, fragrance study in orchid is not as well established as in other flowers such as rose (Guterman et al., 2002), petunia (Verdonk
3. Expressed sequence tags (ESTs)
Expressed Sequence Tags (ESTs) are partial sequences generated from single-pass sequencing (5’- or 3’- end) from a reverse transcription of mRNA representing tissue of interest or a particular developmental stage of an organism (Adam et al., 1991). In plant, EST approach was first used in the model plant
EST libraries for many plant species such as
3.1. Importance of ESTs
EST is a fast, efficient and valuable tool for gene expression, genome annotation and evolutional studies. Analysis of ESTs provides a platform for functional genomics study as well as uncovering the potentially novel genes, and poses an avenue for genome sequencing projects (Ayeh, 2008; Kisiel & Podkowinski, 2005; Li et al., 2010; Lindqvist et al., 2006). Early EST projects were focused mostly on economically important plants and crop plants. In subsequent years, more EST projects on plants yet-to-achieve high economical impact started to materialise. The development of ESTs for some of the plants species from the early years until now is summarised in Table 2.
EST also proves to be a beneficial resource for comparative genomic studies in plant. Hsiao et al. (2006) deduced monoterpene biosynthesis pathway and identified a few fragrance-related genes in
3.2. Discovery of fragrance-related genes from VMPESTs
Expressed sequence tags (ESTs) have been developed from many monocot plant species including
A VMP floral cDNA library was previously constructed from opened flowers at different developmental stages and time-points (Chan et al., 2009). All the cDNA clones with the inserts sizes of 0.5 kb to 1.6 kb were mass excised and single-pass 5’-sequenced. From our attempt, a total of 2,132 ESTs was generated. This VMP dbEST (designated as VMPEST) was clustered, annotated and further classified with Gene Ontology (GO) identifier into three categories: Molecular Functions (51.2%), Cellular Components (16.4%) and Biological Processes (24.6%). Around 3.1% of the VMPEST had hits with other orchid species such as dendrobium, phalaenopsis, oncidium,
From the VMPEST, several fragrance-related transcripts were selected for full-length isolation and expression analysis using real-time quantitative RT-PCR. They were
Among all the transcripts analysed,
4. Simple sequence repeat and its importance
Simple Sequence Repeat (SSR) or microsatellite is a short tandem repeats of a unique DNA sequence with one to six nucleotides motif (Jacob et al., 1991). SSR is a famous molecular marker because of its hyper variability, relative abundance, highly reproducible, multiallelic diversity, co-dominantly inherited and extensive coverage of the genome (Mohan et al., 1997).
Owing to its desirable genetic attributes, SSRs have been utilized in genetic and genomic analyses including genetic mapping, marker assisted plant breeding, development of linkage map, and ecology studies (Kalia et al., 2011; Sonah et al., 2011; Yue et al., 2006). Yue et al. (2006) reported the usage of SSRs in the protection of new
To date, the used of SSRs markers have been reported in several monocot and dicot species including raspberry and blackberry (Stafne et al., 2005), rice (Chakravarthi & Naravaneni, 2006), common bean (Yu et al., 2000),
So far, the reported SSRs generated from vandaceous orchids were used as selective marker only. Phuekvilai et al. (2009) generated SSRs from 33 vandas species for the sole purpose of identifying and evaluating the purity of cultivar in commercial samples. However, the identification and development of SSRs for VMP will be channelled towards facilitating the screening of any potential fragrance-related transcripts from closely related species. Besides, it will be used to determine the extent of inter-species transferability of genes, which had been reported in many plant species (Chapman et al., 2009; Stafne et al., 2005; Wang et al., 2004).
4.1. Data mining of VMPEST-SSR
In recent years, genic microsatellite or EST-SSRs which is less time consuming and relatively easy to develop has replaced the genomics SSRs (Sharma et al., 2007). The publicly available ESTs sequences facilitate the development of SSRs by using the SSR identification tools. These search tools include MISA (MIcroSAtellite), SSRIT (SSR Identification Tool), SciRoKo, TRF (Tandem Repeat Finder), Sputnik, SSRfinder, SAT (SSR Analysis Tool), Poly and SSR Primer. It is deemed important to choose a search tool which is user-friendly and has unlimited access to a non-redundant database (this was well reviewed by Kalia et al., 2011). The first attempt to develop such EST-SSR marker was in rice by Miyao et al. (1996).
SSRIT which is accessible at URL (http://www.gramene.org) was used to identify the SSR motifs in our VMPEST. The script assessed the sequences uploaded in FASTA-formatted files and detected the SSR motifs, the number of repeats as well as identified the sequence corresponded to the SSRs. A total of 98 (9.4%) unigenes containing 112 SSRs with motifs length ranging from two to six nucleotides were detected from VMPEST.
The VMPEST-SSRs were classified into 2 groups with 88.4% belonging to Class I (
Sharma et al. (2009) stressed that the length of repeats and the tools used in the EST-SSRs mining play a significant role in EST-SSR occurrences. The di-nucleotide motif (AT/TA) was present with the most abundance (33.9%) in our VMPEST-SSR. Such observation of A- and T-rich signatures being the most common repeat motifs in the VMPEST-SSRs is also reflected in the early findings of Chagne et al. (2004), Lagercrantz et al. (1993), and Morgante & Olivieri (1993). Interestingly, Blair et al. (2009) found that this motif occurred mainly in the 3’-end of the common bean cDNA clones.
Nevertheless, whatever mined results we obtained from our study, each SSR needs to be validated with further analyses such as selection of SSR primers and screening for utility in vandaceous orchids.
Besides EST-SSR, there are other alternative data mining techniques such as expressed sequence tag-single nucleotide polymorphism (EST-SNP) in maize (Batley et al., 2003), barley
The VMPEST dataset is a potential asset in facilitating the molecular biology and cloning of more genes involved in the fragrance biosynthesis pathway(s). Several fragrance-related transcripts were identified from our VMPEST including
Authors would like to thank the Ministry of Higher Education Malaysia and Universiti Putra Malaysia for financial support through the Fundamental Research Grant Scheme (02-12-10-1002FR) and Research University Grant Scheme (05-04-08-0551RU), respectively.
Aceto S. Gaudio L. 2011The MADS and the beauty: genes involved in the development of orchid flowers. Current Genomics, 12 342 356
Adam M. D. Kelley J. M. Gocayne J. D. Dubnick M. Polymeropoulos M. H. Xiao H. Merril C. R. Wu A. Olde B. Moreno R. F. Kerlavage A. R. Mc Combie W. R. Venter J. C. 1991Complementary DNA sequencing: expressed sequence tags and human genome project. Science, 252 1651 1656
Adelene S. A. L. Janna O. A. Mohd P. Norazizah S. Raha A. R. Puad A. (2012 2012Functional expression of an orchid fragrance gene in Lactococcus lactis. International Journal of Molecular Sciences, 13 1582 1597
Agrawal A. Janssen A. Bruin J. Posthumus M. Sabelis M. 2002An ecological cost of plant defence: attractiveness of bitter cucumber plants to natural enemies of herbivores. Ecological Letters, 5 377 385
Ajjikuttira P. A. Woon M. H. Ryu K. H. Chang C. A. Loh C. S. Wong S. M. Ridge K. 2002Genetic variability in the coat protein genes of two orchid viruses: cymbidium mosaic virus and odontoglossum ringspot virus. Archives of Virology, 147 1943 1954
Alba R. Fei Z. J. Liu Y. Moore S. Debbie P. Cohn J. Ascenzo M. Gordon J. Rose J. Martin G. Tanksley S. Bouzayen M. Jahn M. Giovannoni J. 2004ESTs, cDNA microarrays and gene expression profiling: tools for dissecting plant physiology and development. The Plant Journal, 39 697 714
Allegre M. Argout X. Boccara M. Fouet O. Roguet Y. Berard A. Thevenin J. M. Chauveau A. Rivallan R. Clement D. Courtois B. Gramacho K. Boland-Auge A. Tahi M. Umaharan P. Brunel D. Lanaud C. 2011Discovery and mapping of a new expressed sequence tag-single nucleotide polymorphism and simple sequence repeat panel for large-scale genetic studies and breeding of Theobroma cacao L. DNA Research, doi:dnares/dsr039
Arditti J. 1992Fundamentals of orchid biology, John Wiley & Sons Inc., New York, USA.
Arimura G. Ozawa R. Kugimiya S. Takabayashi J. Bohlmann J. 2004Herbivore-induced defense response in a model legume: two-spotted spider mites, Tetranychusurticae, induce emission of (E)-β-ocimene and transcript accumulation of (E)-β-ocimene synthase in Lotus japonicas. Plant Physiology, 135 1976 1983
Ayeh K. O. 2008Expressed sequence tags (ESTs) and single nucleotide polymorphisms (SNPs): emerging molecular marker tools for improving agronomic traits in plant biotechnology. African Journal of Biotechnology, 7 331 341
Balakrishnan V. P. Vadivel A. 2012In silico RAPD priming sites in expressed sequences and iSCAR markers for oil palm. Comparative and Functional Genomics, doi:
Batley J. Barker G. O’Sullivan H. Edwards K. J. Edwards D. 2003Mining for single nucleotide polymorphisms and insertions/deletions in maize expressed sequence tag data. Plant Physiology, 132 84 91
Baudino S. Caissard-C J. Bergougnoux V. Jullien F. Magnard-L J. Scalliet G. Cock J. M. Hugueney P. 2007Production and emission of volatile compounds by petal cells. Plant Signal Behaviour, 2, 525-526,
Baysal T. Demirdoven A. 2007Lipoxygenase in fruits and vegetables: A review. Enzyme and Microbial Technology, 40 491 496
Beekwilder J. Alvarez-Huerta M. Neef E. Verstappen F. Bouwmeester H. Aharoni A. 2004Functional characterization of enzymes forming volatile esters from strawberry and banana. Plant Physiology, 135 1865 1878
Bick J. A. Lange M. 2003Metabolic cross talk between cytosolic and plastidial pathways of isoprenoids biosynthesis: unidirectional transport of intermediates across the chloroplast envelope membrane. Archives of Biochemistry and Biophysics, 415 146 154
Blair M. W. Torres M. M. Giraldo M. C. Pedraza F. 2009Development and diversity of Andean-derived, gene-based microsatellites for common bean (Phaseolus vulgaris L.). BMC Plant Biology, 9 100 113
Boatright J. Negre F. Chen X. Kish C. Wood B. Peel G. Orlova I. Gang D. Rhodes D. Dudareva N. 2004Understanding in vivo benzenoid metabolism in petunia petal tissue. Plant Physiology, 135 1993 2011
Bory S. Silva D. Risterucci A. Grisoni M. Besse P. Duval M. 2008Development of microsatellite markers in cultivated vanilla: Polymorphism and transferability to other vanilla species. Scientia Horticulturae, 115 420 425
Bradbury L. 2007Identification of the gene responsible for fragrance in rice and characterization of the enzyme transcribed from this gene and its homologs. Doctor of Philosophy dissertation, Southern Cross University, Australia.
Brenner E. D. Stevenson D. W. Mc Combie R. W. Katari M. S. Rudd S. A. Mayer K. F. Palenchar P. M. Runko S. J. Twigg R. W. Dai G. Martienssen R. A. Benfey P. N. Coruzzi G. M. 2003Expressed sequence tag analysis in Cycas, the most primitive living seed plant. Genome Biology, 4 78 88
Chagne D. Chaumeil P. Ramboer A. Collada C. Guevara A. Cervera M. T. Vendramin G. G. Garcia V. Frigerio-M J. Echt C. Richardson T. Plomion C. 2004Cross-species transferability and mapping of genomic and cDNA SSRs in pines. Theoretical and Applied Genetics, 109 1204 1214
Chai D. Yu H. 2007Recent advances in transgenic orchid production. Orchid Science and Biotechnology, 1 34 39
Chakravarthi B. K. Naravaneni R. 2006SSR marker based DNA fingerprinting and diversity study in rice (Oryza sativa L.). African Journal of Biotechnology, 5 684 688
Chan W. S. Janna O. A. Parameswari N. Maziah M. 2009Molecular characterization of a new 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) transcript from Vanda Mimi Palmer. Scientia Horticulturae, 121 378 382
Chan W. S. Janna O. A. Parameswari N. 2011Isolation, cloning and characterization of fragrance-related transcripts from Vanda Mimi Palmer. Scientia Horticulturae, 127 388 397
Chandasekar A. Riju A. Sithara K. Anoop S. Eapen S. J. 2009Identification of single nucleotide polymorphism in ginger using expressed sequence tags. Bioinformation, 4 119 122
Chapman M. A. Hvala J. Strever J. Matvienko M. Kozik A. Michelmore R. W. Tang S. Knapp S. J. Burke J. M. 2009Development, polymorphism and cross-taxon utility of EST-SSR markers from safflower (Carthamus tinctorius L.). Theoretical and Applied Genetics, 120 85 91
Chen M. S. Wang G. J. Wang R. L. Wang J. Song S. Q. Xu Z. F. 2011Analysis of expressed sequence tags from biodiesel plant Jatropha curcas embryos at different developmental stages. Plant Science, doi:10.1016/j.plantsci.2011.08.003
Cherri M. Jullien F. Heizmann P. Baudino S. 2007Fragrance heritability in hybrid tea roses. Scientia Horticulture, 113 177 181
Deleu W. Esteras C. Roig C. Gonzalez-To M. Fernandez-Silva I. Gonzalez-Ibeas D. Blanca J. Aranda M. A. Arus P. Nuez F. Monforte A. J. Pico M. B. Garcia-Mas J. 2009A set of EST-SNPs for map saturation and cultivar identification in melon. BMC Plant Biology, 9, 90, doi:10.1186/1471-2229-9-90
Dudareva N. Cseke L. Blanc V. Pichersky E. 1996Evolution of floral scent in Clarkia: novel patterns of S-linalool synthase gene expression in the C. breweri flower. Plant Cell, 8 1137 1148
Dudareva N. D’Auria J. Nam K. H. Raguso R. Pichersky E. 1998Acetyl-CoA:benzylalcohol acetyltransferase- an enzyme involved in floral scent production in Clarkia breweri. The Plant Journal, 14 297 304
Dudareva N. Pichersky E. Gershenzon J. 2004Biochemistry of plant volatiles. Plant Physiology, 135 1839 1902
Eun A. J. Wong S. 2000Molecular beacons: a new approach to plant virus detection. Phytopathology, 90 269 275
Eun A. J. Huang L. Chew F. Li S. F. Wong S. 2002Detection of two orchid viruses using Quartz crytal microbalance-based DNA biosensors. Phytopathology, 92 654 658
Fernandes J. Brendel V. Gai X. Lal S. Chandler V. L. Elumalai R. P. Galbraith D. W. Pierson E. A. Walbot V. 2002Comparison of RNA expression profiles based on maize expressed sequence tag frequency analysis and micro-array hybridization. Plant Physiology, 128 896 910
Frowine S. A. 2005Fragrant orchids, Timber Press Inc., Oregon, USA.
Go R. Yong W. S. Y. Unggang J. Salleh R. 2010Orchids of Perlis, Jewels in the Forests, Revised Edition. Perlis Forestry Department and Universiti Putra Malaysia Press, Kuala Lumpur, Malaysia, 152
Go R. Khor H. E. Tan M. C. Janna O. A. Farah Alia. N. Ng Y. J. Muskhazli M. Mohd Nazre. S. Rosimah N. 2012Current knowledge on orchid diversity in Peninsular Malaysia including new records and discovery. (Manuscript submitted and under review)
Gonzalez-Mas M. C. Garcia-Riano L. M. Alfaro C. Rambla J. L. Padilla A. I. Gutierrez A. (2008 2008Headspace-based techniques to identify the principal volatile compounds in red grape cultivars. International Journal of Food Science & Technology, 44 510 518
Guterman I. Shalit M. Menda N. Piestun D. Dafny Y. Shalev G. Bar E. Davydov O. Ovadis M. Emanuel M. Wang J. Adam Z. Pichersky E. Lewinsohn E. Zamir D. Vainstein A. Weiss D. 2002Rose scent: genomics approach to discovering novel floral fragrance-related genes. The Plant Cell, 14 2325 2338
Hofte H. Desprez T. Amselem J. Chiapello H. Rouze P. Caboche M. Moisan A. Jourjon M. F. Charpenteau J. L. Berthomieu P. Guerrier D. Giraudat J. Quigley F. Thomas F. Yu D. Y. Raynal M. Cooke R. Grellet F. Marcillac P. Gigot C. Fleck J. Phillips G. Axelos M. Bardet C. Tremousaygue D. Lescure B. 1994An inventory of 1152 expressed sequence tags obtained by partial sequencing of cDNAs from Arabidopsis thaliana. Plant Journal, 4 1051 1061
Hsiao Y. Tsai W. Kuoh C. Huang T. Wang H. Wu T. Leu Y. Chen W. Chen H. 2006Comparison of transcripts in Phalaenopsis bellina and Phalaenopsis equestris (Orchidaceae) flowers to deduce monoterpene biosynthesis pathway. BMC Plant Biology, 6 14 27
Hsiao Y. Y. Pan Z. J. Hsu C. C. Yang Y. P. Hsu Y. C. Chuang Y. C. Shih H. H. Chen W. H. Tsai W. C. Chen H. H. 2011Research on Orchid Biology and Biotechnology. Plant and Cell Physiology, 52(9), 1467-1486.
Hsu C. C. Chung Y. L. Chen T. C. Lee Y. L. Kuo Y. T. Tsai W. C. Hsiao Y. Y. Chen Y. W. Wu W. L. Chen H. H. 2011An overview of the Phalaenopsis orchid genome through BAC end sequence analysis. BMC Plant Biology, 11 3 13
Hsu H. T. Vongsasitorn D. Lawson R. H. 1992An improved method for serological detection of cymbidium mosaic potexvirus infection in orchids. Phytopathology, 82 491 495
Hu W. W. Wong S. M. 1998The use o DIG-labelled cRNA probes for the detection of cymbidium mosaic potexvirus (CymMV) and odontoglossum ringspot tobamovirus (ORSV) in orchids. Journal of Virological Methods, 70 193 199
Huang Y. Li F. Chen K. 2010Analysis of diversity and relationships among Chinese orchid cultivars using EST-SSR markers. Biochemical Systematics and Ecology, 38 93 102
Jacob H. J. Lindpaintner K. Lincoln S. E. Kusumi K. Bunker R. K. Mao Y. P. Ganten D. Dzau V. J. Lander E. S. 1991Genetic mapping of a gene causing hypertensive rat. Cell, 67 213 224
Janna O. A. Tan L. Aziz A. Maziah M. Puad A. Azlan J. 2005Dull to behold, good to smell… Vanda Mimi Palmer. BioTech Communications, Faculty of Biotechnology & Biomolecular Science Research Bulletin, Universiti Putra Malaysia, Malaysia, 1 21 25 1823-3279
Jiang D. Y. E. Q. L. Wang F. S. Cao L. 2010The mining of citrus EST-SNP and its application in cultivar discrimination. Agricultural Sciences in China, 9 179 190
Kalia R. K. Rai M. K. Kalia S. Singh R. Dhawan A. K. 2011Microsatellite markers: an overview of the recent progress in plants. Euphytica, 177 309 334
Khentry Y. Paradornuwat A. Tantiwiwat S. Phransiri S. Thaveechai N. 2006Incidence of cymbidium mosaic virus and odontoglossum ringpot virus in Dendrobium spp. in Thailand. Crop Protection, 25 926 932
Kishor R. Sha Valli. Khan P. S. Sharma G. J. (2006 2006Hybridization and in vitro culture of an orchid hybrid Ascocenda ‘Kangla’. Scientia Horticulturae, 108 66 73
Kisiel A. Podkowinski J. 2005Expressed Sequence Tags and their applications for plants research. Acta Physiologiae Plantarum, 27 157 161
Kishor R. Devi H. Jeyaram K. Singh M. 2008Molecular characterization of reciprocal crosses of Aerides vandarum and Vanda stangeana (Orchidaceae) at the protocorm stage. Plant Biotechnology Reports, 2 145 152
Lagercrantz U. Ellegren H. Anderson L. 1993The abundance of various polymorphic microsatellite motifs differs between plants and vertebrates. Nucleic Acids Research, 21 1111 1115
Lavid N. Wang J. Shalit M. Guterman I. Bar E. Beuerle T. Menda N. Shafir S. Zamir D. Adam Z. Vainstein A. Weiss D. Pichersky E. Lewinsohn E. 2002O-methyltransferases involved in the biosynthesis of volatile phenolic derivatives in rose petals. Plant Physiology, 129 1899 1907
Leonard J. Watson C. Carter A. Hansen J. Zemetra R. Santra D. Campbell K. Riera-Lizarazu O. 2008Identification of a candidate gene for the wheat endopeptidaseEp-D1 locus and two other STS markers linked to the eyespot resistance gene Psh1. Theoretical & Applied Genetics, 116 261 270
Li X. Shangguan L. Song C. Wang C. Gao Z. Yu H. Fang J. 2010Analysis of expressed sequence tags from Prunus mume flower and fruit and development of simple sequence repeat markers. BMC Genetics, 11 66 76
Lindqvist C. Scheen A. Yoo M. Grey P. Oppenheimer D. G. Leebens-Mack J. H. Soltis D. E. Soltis P. S. Albert V. A. 2006An expressed sequence tag (EST) library from developing fruits of an Hawaiian endemic mint (Stenogyne rugosa, Lamiaceae): characterization and microsatellite markers. BMC Plant Biology, 6 16 30
Low E. T. Alias H. Boon S. H. Shariff E. Tan C. Y. Ooi L. Cheah S. C. Raha A. R. Wan K. L. Singh R. 2008Oil palm (Elaeis guineensis Jacq.) tissue culture ESTs: Identifying genes associated with callogenesis and embryogenesis. BMC Plant Biology, 8 62 81
Majetic C. Raguso R. Ashman T. 2009The sweet smell of success: floral scent affects pollinators attraction and seed fitness in Hesperis matronalis. Functional Ecology, 23 480 487
Micheneau C. Fournel J. Gaucin-Bialecki A. Pailler T. (2008 2008Auto-pollination in a long-spurred endemic orchid (Jumellea stenophylla) on Reunion Island (Mascarene Archipelago, Indian Ocean). Plant Systematics and Evolution, 272 11 22
Miyao A. Zhong H. Monna L. Yano M. Yamamoto K. Havukkala I. Minobe Y. 1996Characterization and genetic mapping of simple sequence repeats in rice genome. DNA Research, 3 233 238
Mohan M. Nair S. Bhagwat A. Krishna T. G. Yano M. Bhatia C. R. Sasaki T. 1997Genome mapping, molecular markers and marker-assisted selection in crop plants. Molecular Breeding, 3 87 103
Mohd-Hairul A. R. Parameswari N. Ee Gwendoline C. L. Janna O. A. 2010Terpenoid, benzenoid and phenylpropanoid compounds in the floral scent of Vanda Mimi Palmer. Journal of Plant Biology, 53 358 366
Morgante M. Olivieri A. M. 1993PCR-amplified microsatellites as markers in plant genetics. Plant Journal, 3 175 182
Nagaraj S. Gasser R. Ranganathan S. 2006A hitchhiker’s guide to expressed sequence tag (EST) analysis. Briefing in Bioinformatics, 8 6 21
Naji A. M. Moghaddam M. Ghaffari M. R. Irandoost H. P. Farsad L. K. Pirseyedi S. M. Mohammadim S. A. Ghareyazie B. Mardi M. 2008Validation of EST-derived STS markers localized on Qfhs.ndsu-3BS for Fusarium head blight resistance in wheat using a ‘Wangshuibai’ derived population. Journal of Genetics & Genomics, 35 625 628
New Comb. R. D. Crowhurst R. N. Gleave A. P. Rikkerink E. Allan A. C. Beuning L. L. Bowen J. H. Gera E. Jamieson K. R. Janssen B. J. Laing W. A. Mc Artney S. Nain B. Ross G. S. Snowden K. C. Souleyre E. Walton E. F. Yauk Y. K. 2006Analyses of expressed sequence tags from apple. Plant Physiology, 141 147 166
Nojima S. Kiemle D. J. Webster F. X. Apperson C. S. Schal C. 2011Nanogram-scale preparation and NMR analysis for mass-limited small volatile compounds. PLoS ONE, 6: e18178. Doi:10.1371/journal.pone.0018178
Ooi J. M. 2005Screening of a random peptide library with CymMV for potential development of diagnostic kits. Master dissertation, Malaysia University of Science and Technology, Selangor Darul Ehsan, Malaysia.
Park Y. S. Jeong M. H. Lee S. H. Moon J. S. Cha J. S. Kim H. Y. Cho T. J. 2005Activation of defense responses in chinese cabbage by a nonhost pathogen, Pseudomonas syringae pv. tomato. Journal of Biochemistry and Molecular Biology, 38 748 754
Phuekvilai P. Pongtongkam P. Peyachoknagul S. 2009Development of microsatellite markers for Vanda orchid. Natural Science, 43 497 506
Pichersky E. Dudareva N. 2007Scent engineering: toward the goal of controlling how flowers smell. TRENDS in Biotechnology, 25 105 110
Pichersky E. Noel J. Durareva N. 2006Biosynthesis of plant volatiles: nature’s diversity and ingenuity. Science, 311 808 811
Pinheiro F. Santos M. O. Barros F. Meyer D. Salatino A. Souza A. P. Cozzolino S. 2008Isolation and characterization of microsatellite loci in the Brazilian orchid Epidendrum fulgens. Conservation Genetics, 9 1661 1663
Rada F. Jaimez R. 1992Comparative ecophysiology and anatomy of terrestrial and epiphytic Anthurium bredemeyeri Schott in a tropical Andean cloud forest. Journal of Experimental Botany, 43 723 727
Raguso R. 2008Wake up and smell the roses: The ecology and evolution of floral scent. Annual Review of Ecology, Evolution and Systematics, 39 549 569
Rani P. Pant R. Jain R. 2010Serological detection of cymbidium mosaic and odontoglossumringspot viruses in orchids with polyclonal antibodies produced against their recombinant coat proteins. Journal of Phytopathology, 158 542 545
Rudd S. 2003Expressed sequence tags:alternative or complement to whole genome sequences? TRENDS IN Plant Science, 8 321 328
Schiestl F. P. 2010The evolution of floral scent and insect chemical communication. Ecology Letters, 13 643 656
Sharma P. Grover A. Kahl G. 2007Mining microsatellites in eukaryotic genomes.TRENDS in Biotechnology, 25 490 498
Sharma R. Bhardwaj P. Negi R. Mohapatra T. Ahuja P. 2009Identification, characterization and utilization of unigenes derived microsatellite markers in tea (Camellia sinensis L.). BMC Plant Biology, 9 53 76
Sherpa A. Hallan V. Pathak P. Zaidi A. 2007Complete nucleotide sequence analysis of cymbidium mosaic virus Indian isolate: further evidence for natural recombination among potexviruses. Journal of Bioscience. 32 663 669
Shuttleworth A. Johnson S. 2009A key role for floral scent in a wasp-pollination system in Eucomis (Hyacinthaceae). Annals of Botany, 103 715 725
Sonah H. Deshmukh R. K. Sharma A. Singh V. P. Gupta D. K. Gacche R. N. Rana J. C. Singh N. K. Sharma T. R. 2011Genome-wide distribution and organization of microsatellite in plants: an insight into marker development in Brachypodium. PLoS ONE, 6 1 9
Song Q. Jia G. Zhu Y. Grant D. Nelson R. Hwang-Y E. Hyten D. Cregan P. 2010Abundance of SSR motifs and development of candidate polymorphic SSR markers (BARCSOYSSR_1.0) in soybean. Crop Science, 50 1950 1960
Spitcer B. Ben Zvi. M. Ovadis M. Marhevla E. Barkai O. Edelbaum O. Marton I. Masci T. Alon M. Morin S. Rogachev I. Aharoni A. Vainstein A. 2007Reverse genetics of floral scent: application of tobacco rattle virus-based gene silencing in petunia. Plant Physiology, 145 1241 1250
Stafne E. T. Clark J. R. Weber C. A. Graham J. Lewers K. S. 2005Simple sequence repeat (SSR) markers for genetic mapping of raspberry and blackberry. Journal of American Horticultural Science, 130 1 7
Teh S. L. 2011Development of a floral expressed sequence tags resource from and characterization of fragrance-related gene transcripts in Vanda Mimi Palmer. Master dissertation, Universiti Putra Malaysia, Selangor Darul Ehsan, Malaysia.
Teh S. L. Chan W. S. Janna O. A. Parameswari N. 2011Development of expressed sequence tag resources for Vanda Mimi Palmer and data mining for EST-SSR. Molecular Biology Reports, 38 3903 3909
Temnykh S. De Clerck G. Lukashova A. Lipovich L. Cartinhour S. Mc Couch S. 2001Computational and experimental analysis of microsatellites in rice (Oryza sativa L.): frequency, length variation, transposon associations and genetic marker potential. Genome Research, 1441 1452
Teo C. K. H. 1981Tropical orchid hybrids, FEP International Sdn. Bhd., Singapore.
Teoh E. S. 1980Asian orchids, Times Books International, Singapore.
Terp N. Gobel C. Brandt A. Feussner I. 2006Lipoxygenases during Brassica napus seed germination. Phytochemistry, 67 2030 2040
Tsai W. Hsiao Y. Lee S. Tung C. Wang D. Wang H. Chen W. Chen H. 2006Expression analysis of the ESTs derived from the flower buds of Phalaenopsis equestris. Plant Science, 170 426 432
van Moerkercke A. Galvan-Ampudia C. S. Verdonk J. C. Haring M. A. Schuurink R. C. 2012Regulators of floral fragrance production and their target genes in petunia are not exclusively active in the epidermal cells of petals. Journal of Experimental Botany, doi:10.1093/jxb/ers034
van Tunen A. J. Eikelboom W. Angenent G. C. 1993Floral organogenesis in Tulipa. Flowering Newsletter, 16 33 37
Vainstein A. Lewinsohn E. Pichersky E. Weiss D. 2001Floral fragrance.New inroads into an old commodity. Plant Physiology, 127 1383 1389
Varshney R. K. Chabane K. Hendre P. S. Aggarwal R. K. Graner A. 2007Comparative assessment of EST-SSR, EST-SNP and AFLP markers fir evaluation of genetic diversity and conservation of genetic resources using wild, cultivated and elite barleys. Plant Science, 173 638 649
Vejaratpimol R. Channuntapipat C. Pewnim T. Ito K. Iizuka M. Minamiura N. 1999Detection and serological relationship of cymbidium mosaic potexvirus isolates. Journal of Bioscience and Bioengineering, 87 161 168
Verdonk J. C. Haring M. A. van Tunen A. J. Schuurink R. C. 2005ODORANT1 regulates fragrance biosynthesis in petunia flowers. The Plant Cell, 17 1612 1624
Wang M. L. Gillaspie A. G. Newman M. L. Dean R. E. Pittman R. N. Morris J. B. Pederson G. A. 2004Transfer of simple sequence repeat (SSR) markers across the legume family for germplasm characterization and evaluation. Plant Genetic Resource, 2 107 119
Yu K. Park S. J. Poysa V. Gepts P. 2000Integration of simple sequence repeat (SSR) markers into a molecular linkage map of common bean (Phaseolus vulgaris L.). The American Genetic Association, 91 429 434
Yue G. H. Lam-Chan L. T. Hong Y. (2006 2006Development of simple sequence repeat (SSR) markers and their use in identification of Dendrobium varieties. Molecular Ecology Resources, 6 832 834
Zhang D. Choi D. Wanamaker S. Fenton R. Chin A. Malatrasi M. Turuspekov Y. Walia H. Akhunov E. Kianian P. Otto C. Deal K. Echenique V. Stamova B. Ross K. Butler G. Strader L. Verhey S. Johnson R. Altenbach S. Kothari K. Tanaka C. Shah M. Laudencia-Chingcuanco D. Han P. Miller R. Crossman C. Chao S. Lazo G. Klueva N. Gustafson J. Kianian S. Dubcovsky J. Walker-Simmons P. Gill K. Dvorak J. Anderson O. Sorrells M. Mc Guire P. Qualset C. Nguyen H. Close T. 2004Construction and evaluation of cDNA libraries for large-scale expressed sequence tag sequencing in wheat (Triticum aestivum L.). Genetics, 168 595 608