Some of the recent studies with plant bioassays.
Abstract
Plants are present in almost all areas of the world and can accumulate many chemical compounds present in the soil, water, and atmosphere. As these chemicals which are potentially mutagenic or carcinogenic are absorbed by the plants sharing the same environment with us, bioassays on plants can be used to detect the presence of environmental hazards. Another reason for selecting plants for assessing adverse effects of these chemicals is the ease of experimentation with plants. Evaluating the effect of a substance on basic plant characteristics such as growth, survival, or reproduction is straightforward and repeatable. Thus, various plant species are commonly utilized as indicators of adverse environmental conditions. This chapter covers the detection of environmental mutagens through plant bioassays, considering the increasing importance of biomonitoring using plants for assessing the mutagenicity of relevant chemicals and industrial waste. From this point of view, a detailed literature search was made on the subject. The genotoxic, cytotoxic, and molecular studies have been investigated and the most useful and important parts and key points of these methods were summarized. This review would be useful for scientists who are planning to conduct research on plant bioassays with different types of methods and chemicals.
Keywords
- ecotoxicology
- cytotoxicity
- genotoxicity
- phytotoxicity
- mutagens
1. Introduction
Plants are the essential elements of agriculture and forestry and maintain the healthy environment for the rest of the species by producing oxygen and organic carbon compounds. Higher plants are preeminent indicators of genotoxic effects caused by chemical substances existing in the environment and therefore be utilized for detecting environmental mutagens [1]. They are exposed to many stress factors including chemical compounds and radiation affecting their seed germination, seedling growth, and floral and fruit development. These stress factors can adversely affect the quality and quantity of the product with leading to morphological, anatomical, physiological, biochemical, and molecular damage to plants [2]. There are different kinds of methods for examining phytotoxicity and genotoxicity because usually there is no standard national procedure. Therefore, the parameters of these methods vary depending on the test substances, the test plants, or the individual procedures. Because of its simplicity, low cost, and relatively high sensitivity, application of plant bioassays is usually favored over other available systems in discovering adverse effects caused by chemical substances, or pollution, existing in the environment [3]. Despite these benefits described above, there are also some limitations in using plant bioassays, such as the longer life span of plants than
Plant species | Test substance | Method | Reference |
---|---|---|---|
Wastewater | Micronucleus method | Liu et al. [9] | |
Pesticide | Micronucleus and stamen hair bioassays | Fadic et al. [10] | |
Aniline | Micronucleus, mitotic index, and chromosomal aberration | Tao et al. [11] | |
Insecticide | Sister chromatid exchange | Quintana et al. [12] | |
Mercury | Real-time PCR FISH | Zhen et al. [13] | |
Ionizing radiation | TUNEL test | Scaldaferro et al. [14] | |
Volatile organic compounds | Comet assay | Naroi-et et al. [15] | |
Lead stress | RAPD-PCR | Venkatachalam et al. [16] |
Plant bioassays are usually based on the detection of chromosomal abnormalities in mitosis, sister chromatid exchanges (SCEs), and, recently, on the DNA damage analysis. Point mutations such as chlorophyll mutations in leaves, waxy mutations, or embryo mutations of
2. Seed germination and root elongation tests
Many plant species have been recommended for ecotoxicity tests using seed germination and root elongation methods. Among them, cabbage, lettuce, and oats are recommended by the US Environmental Protection Agency (EPA) (1983) [18], the Organization for Economic Cooperation and Development (OECD) (1984) [19], and the Food and Drug Administration (FDA) (1987) [20]. Carrot, cucumber, and tomato are also suggested by the EPA and FDA, wheat is accepted by the FDA and OECD, and rice is also mentioned by the OECD. Although not mentioned in any of these documents, millet has been studied at the Illinois State Water Survey for several years [21]. Most frequently used species are
3. Cytogenetic techniques
The frequency and the type of chromosome abnormalities in different phases of mitosis and the micronuclei frequency of interphase cells are analyzed by cytogenetic tests. The DNA damage caused by the genotoxic agents could either be repaired or otherwise could be lead to the DNA alterations. Chromosome abnormalities are the results of DNA double-strand breaks that were unrepaired or inaccurately repaired. Chromosomes are rearranged since broken chromosome ends become “sticky” and may combine with other broken chromosome ends. After mutagenic treatment, because of the chromosomal rearrangements and acentric fragments, dicentric bridges could be observed in mitotic cells of the first cell cycle. Micronuclei frequency also decreases in the interphase cell in the next cell cycle [36]. The micronucleus (MN) test,
3.1. Allium /Vicia chromosome aberration test
Several mutagens can be detected cytologically by cellular inhibition; disruption in metaphase; induction of chromosomal aberrations, numerical and structural, ranging from chromosomal fragmentation to the disorganization of the mitotic spindle; and consequently all subsequent dependent mitotic phases. The microscopic analysis includes mitotic index, micronuclei presence in interphase cells, and chromosomal aberrations in late anaphase and early telophase cells score. Approximately 1000 cells from all the stages of dividing cells in mitosis are counted in order to find the mitotic index value. Chromosomal abnormalities can be determined, and then, they are scored in the first 100 cells in different stages of mitotic division. The mostly used method to determine all of the abnormalities is to scan the slides from right to left, up, and down [39]. The
3.2. Tradescantia stamen hair mutation and micronucleus analysis
The genus
Ma and Grant [43] have prepared a historical perspective, detailing the importance of this plant in mutation studies. Firstly, the heterozygosity for flower color in
3.3. Sister chromatid exchange
The sister chromatid exchange (SCE) test is developed from the semiconservative DNA replication model which we could see the separation of DNA. The cytogenetic monitoring of exposure to potential mutagens in the environment could be done by SCE which is a highly sensitive cytogenetic tool for detecting DNA damage. It involves firstly the breakage of both DNA strand and then an exchange of whole DNA duplexes. The symmetrical exchange during S phase at one locus between sister chromatids that does not alter chromosome length and genetic information is defined. Taylor was the first scientist who made the SCE test visualized for plant cells, but he used tritium and autoradiography, which provided poor spatial resolution [56]. After Taylor, it was discovered that sister chromatids could be differentiated and revealed SCEs in combination with Hoechst dye 33258 incorporation of the DNA base analog 5′-bromodeoxyuridine (BrdUrd) staining [57]. BrdUrd is a synthetic nucleoside that is an analog of thymidine and is actively incorporated into the newly synthesized DNA during replication process. It is commonly used in the detection of dividing cells in living organisms during the S phase of the cell cycle substituting for thymidine. The standard fluorescence plus Giemsa (FPG) staining method also will enable visualization of SCEs in metaphase spreads of growing cells in medium containing BrdUrd with a light microscope [56]. The frequency of SCEs per chromosome set increases after treatment with genotoxic agents. SCE method was first applied in mammalian cells, and later, it has been shown that it can be applied in plant cells.
Especially plant species that have relatively large and a low number of chromosomes such as
4. Molecular techniques
4.1. Fluorescent in situ hybridization
The classical cytogenetic techniques were usually used for detecting the changes in chromosomal number and morphology. However, chromosome staining with the traditional methods such as Feulgen or orcein staining can fail in the analysis of small changes in chromosome structure. The fluorescent in situ hybridization (FISH) allows the detection and a more detailed localization of chromosomal rearrangements, both in interphase and mitotic nuclei, which gives new possibilities to study chromosomal aberrations [61]. Additionally, it helps to reveal the mechanisms of the formation of chromosomal abnormalities in plant mutagenesis. Although there are a few number of DNA probes for particular plant chromosomes,
4.2. TUNEL test
Another test used to identify apoptosis that has found application in plant genotoxicity studies is the terminal deoxyribonucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) test [64]. TUNEL assay detects DNA fragmentation by the help of fluorescence microscope. TUNEL test is used to detect DNA damage associated with nonapoptotic events such as necrotic cell death induced by exposure to genotoxic chemicals. It is not limited to the detection of apoptotic cells [65] and has also ability to stain cells going through active DNA repair [66]. The regulated cell death plays an important role during development of plants, and it is also essential for plant-specific responses to biotic and abiotic stress factors. The terminal deoxynucleotidyl transferase catalyzes the polymerization of labeled nucleotides to DNA strand breaks in situ. For TUNEL test, successive hand-cut sections of each axis of embedded plant material are stained with propidium iodide (PI) in order to stain the nuclei of dead cells to red and DAPI (4',6-diamidino-2-phenylindol) which can pass through the normal cell membrane and stains the nuclei to blue. DAPI can be used to stain both live and fixed cells. The detection of DNA breaks at a single nucleus can be achieved with TUNEL test within a short time, and the screening of labeled nuclei is easier than other methods. It is recommended for the preliminary genotoxicity investigation of the new identified chemicals [67].
4.3. Single-cell gel electrophoresis (comet assay)
DNA damage in higher plant cells was evaluated by the frequency of chromosomal aberrations in metaphase chromosomes, abnormal anaphase and telophases, and micronuclei; however, these tests measure unrepaired genome damage in cells which have reached mitosis. DNA damage may be originated from DNA metabolism spontaneously or from the effects of environmental factors. There are different kinds and levels of DNA repair mechanisms in cell nucleus to prevent these damages. When the repair mechanisms are ineffective or there was a heavy DNA damage, it may lead to the inhibition of replication, transcription, or protein synthesis; however, in the long term, chromosomal abnormalities or mutations could be formed. It is a sensitive and fast fluorescent technique, which is used to determine the amount of DNA damage on single cell level. After its introduction as “alkaline comet assay,” it has been developed with many modifications for investigating the process of apoptosis and became a workable technique for detecting a variety of DNA damages in plant cells. It allows the determination of double- and single-stranded DNA breaks in a single cell and also helps to measure the level of the migration of DNA by using horizontal gel electrophoresis system [68]. The length of the tail and the amount of the DNA in the head and in the tail are measured to assess the toxicity in a computerized image analysis system. The tail moment (TM) can be calculated to show DNA damage [69]. The comet assay allows fast detection of DNA damage, shortly after the injury, before DNA is repaired, and without any need to wait for progression into mitosis [70]. The presence of a cell wall and the absence of free cells in plant tissues cause technical difficulties for performing the comet assay. Over the past few years, many scientists have improved the methodology for the comet assay on plant cells. Navarrete et al. [70]. developed a simple and efficient mechanical extraction to isolate cell nuclei to overcome these problems. This technique was then improved by Gichner. The different internal parameters such as nucleus isolation methods, filtration and lysis steps, agarose concentration, and the external parameters such as room temperature and light intensity were evaluated during these studies [71].
4.4. Random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) technique
RAPD-PCR is a PCR-based and quite reproducible technique that yields information on a large number of markers without having to obtain DNA sequence information for primer design [72]. Many scientists used RAPD-PCR technique commonly for a variety of purposes such as cultivar identification, genetic diversity assessment, and the construction of phylogenetic relationships [73], and it has been successfully utilized in genotoxicity identification of toxic chemicals. A number of selective and sensitive assays for DNA analysis in ecotoxicology have been developed with the improvement of recent molecular biology techniques. DNA-based techniques such as RFLP, QTL, RAPD, AFLP, SSR, and VNTR are being used to investigate the variations at the DNA sequence level. RAPD-PCR can be used to detect genotoxicity, and differences in RAPD profiles can clearly be shown when comparing DNA fingerprints from untreated and treated individuals to genotoxic agents [74]. Many studies support the view that the RAPD analysis is a highly sensitive method for the detection of DNA damage induced by environmental pollutants like toxic chemicals. RAPD markers are at this moment low valuable markers due to the lack of repeatability. A few work is usually published at this moment using this kind of markers. This kind of study using other DNA markers will be of much more interest.
4.5. Real-time polymerase chain reaction (RT-PCR) technique
Plants have risk of DNA damage due to continuous exposure to environmental mutagens, and thus a variety of repair mechanisms should operate to maintain genome integrity.
5. Flow cytometry
Flow cytometry (FCM) is a rapid and multiparametric technique that theoretically has the potential to detect minute variations in nuclear DNA (nDNA) content, as well as chromosomal damage, in exposed organisms. It can also provide information on polyploidization and evaluate cell cycle dynamics in plants. Pfosser et al. [79]. evaluate the sensitivity of FCM by detecting the variations in DNA content as small as 1% in aneuploid wheat-rye lines. Relatively to DNA damage, Rayburn and Wetzel correlated the coefficient of variation of the G0/G1 peak with chromosomal aberration in aluminum-exposed plants, as this parameter is able to detect broken and rearranged chromosomes in daughter cells [80]. Monteiro et al. also detected an increase in the full peak coefficient of variation (FPCV) of the G0/G1 peak of lettuce plants exposed to Cd [81].
6. Conclusion
Hundreds of new industrial chemicals have been continuously produced to facilitate our lives, and we are not able to be aware of their damage before we investigate their effects on organisms. Plant bioassays serve as a tool to demonstrate the cytotoxic and genotoxic effects of environmental pollutants by means of clear-cut evidence of chromosome damage and gene mutation. These studies could also be useful to establish a database for environmental conditions in the various regions of the world. Some of these simple and clear-cut indicators revealed by plant bioassays could also be used to demonstrate the genotoxic effects of environmental pollution to the general public. The kind of education that is required is not only about teaching people how to detect and eliminate pollutants but also to educate the general public on the root cause of pollution problems. Pollution is related to every facet of human life, and it is life itself that generates pollution. Regulations and guidelines are essential to cure the symptoms of pollution. Plant bioassay studies deserve to be included by the enforcement agencies, particularly of the developing countries, for their regular monitoring of pollution sites.
References
- 1.
Grant WF. The present status of higher plant bioassays for the detection of environmental mutagens. Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis. 1994; 310 (2):175-185. DOI: 10.1016/0027-5107(94)90112-0 - 2.
Büyük İ, Soydam-Aydın S, Aras S. Molecular responses of plants to stress conditions. Türk Hijyen ve Deneysel Biyoloji Dergisi. 2012; 69 (2):97-110. DOI: 10.5505/TurkHijyen.2012.40316 - 3.
Constantin MJ, Owens ET. Introduction and perspective of plant genetic and cytogenetic assays. Mutation Research. 1982; 99 :37-49 - 4.
Grant WF, Salamone MF. Comparative mutagenicity of chemicals selected for test in the International Program on Chemical Safety`s collaborative study on plant systems for the detection of environmental mutagens. Mutation Research. 1994; 310 :187-209 - 5.
Ma TH. The international program on plant bioassays collaborative studies with plant systems. Mutation Research. 1999; 426 :97-98 - 6.
Knasmuller S, Gottmann E, Steinkellner H, Fomin A, Pickl C, Paschke A, God R, Kundi M. Detection of genotoxic effects of heavy metal contaminated soils with plant bioassays. Mutation Research. 1998; 420 :37-48 - 7.
Gopalan HNB. Ecosystem health and human wellbeing: the mission of the International Program on Plant Bioassays. Mutation Research. 1999; 426 :99-102 - 8.
Sandhu SS, de Serres FJ, Gopalan HNB, Grant WF, Velemisky J, Becking GC. An introduction and study design. Mutation Research. 1994; 10 :169-173 - 9.
Liu Y, Liu J, Zhang A, Liu Z. Treatment effects and genotoxicity relevance of the toxic organic pollutants in semi-coking wastewater by combined treatment process. Environmental Pollution. 2017; 220 (Pt A);13-19. DOI: 10.1016/j.envpol.2016.04.095 - 10.
Fadic X, Placencia F, Domínguez AM, Cereceda-Balic F. Tradescantia as a biomonitor for pesticide genotoxicity evaluation of iprodione, carbaryl, dimethoate and 4,4′-DDE. The Science of the Total Environment. 2017; 575 :146-151. DOI: 10.1016/j.scitotenv.2016.09.198 - 11.
Tao N, Liu G, Bai L, Tang L, Guo C. Genotoxicity and growth inhibition effects of aniline on wheat. Chemosphere. 2017; 169 :467-473. DOI: 10.1016/j.chemosphere.2016.11.063 - 12.
Quintana VR, Gómez-Arroyo S, Sánchez-Alarcón J, Milić M, Olivares JL, Waliszewski SM, Cortés-Eslava J, Villalobos-Pietrini R, Calderón-Segura ME. Genotoxic effects of the carbamate insecticide Pirimor-50® in Vicia faba root tip meristems and human lymphocyte culture after direct application and treatment with its metabolic extracts. Arhiv za Higijenu Rada i Toksikologiju. 2016: 67 (4):266-276. DOI: 10.1515/aiht-2016-67-2809 - 13.
Zhen C, Moshun C, Ming J. Hydrogen sulfide alleviates mercury toxicity by sequestering it in roots or regulating reactive oxygen species productions in rice seedlings. Plant Physiology and Biochemistry. 2017; 111 :179-192 - 14.
Scaldaferro MA, Prina AR, Moscone EA, Kwasniewska J. Effects of ionizing radiation on Capsicum baccatum var. pendulum (Solanaceae). Applied Radiation and Isotopes. 2013;79 :103-108. DOI: 10.1016/j.apradiso.2013.04.004 - 15.
Na roi-et V, Chiemchaisri W & Chiemchaisri C. Genotoxicity assessment of volatile organic compounds in landfill gas emission using comet assay in higher terrestrial plant. Bulletin of Environmental Contamination and Toxicology. 2016; 98 :283. DOI: 10.1007/ s00128-016-2012-3 - 16.
Venkatachalam P, Jayalakshmi N, Geetha N, Sahi SV, Sharma NC, Rene ER, Sarkar SK, Favas PJC. Accumulation efficiency, genotoxicity and antioxidant defense mechanisms in medicinal plant Acalypha indica L. under lead stress. Chemosphere. 2017; 171 :544-553 - 17.
Uhl M, Plewa M, Majer BJ, Knasmuller S. Basic principles of genetic toxicology with an emphasis on plant bioassays. In: Maluszynska J, Plewa M, editors. Bioassays in Plant Cells for Improvement of Ecosystem and Human Health. Katowice: Wydawnictwo Uniwersytetu Śląskiego; 2003. pp. 11-30 - 18.
Ratsch HC. Interlaboratory Root Elongation Testing of Toxic Substances on Selected Plant Species. EPA-600/S3-83-051. Corvallis, OR: U.S. Environmental protection Agency; 1983 - 19.
Organization for Economic Cooperation and Development. Terrestrial Plants: Growth test. Paris: OECD guideline for Testing Chemicals; 1984. No: 208 - 20.
Food and Drug Administration. Seed Germination and Root Elongation. Environmental Assessment Technical Assistance Document 4.06, the Center for Food Safety and Applied Nutrition and the Center for Veterinary Medicine. Washington, D.C.: U.S. Department of Health and Human Services; 1987 - 21.
Wang W, Keturi PH. Comparative seed germination tests using ten plant species for toxicity assessment of a metal engraving effluent sample. Water, Air, & Soil Pollution. 1990; 52 :369-376. DOI: 10.1007/BF00229444 - 22.
Pfleeger T, Mc Farlane C, Sherman R, Volk G. A short-term bioassay for whole plant toxicity. In: Gorsuch JW, Lower WR, Wang W, Lewis MA. editors. Plants for Toxicity Assessment. Vol. 2 (ASTM STP 1115). Philadelphia, PA: American Society for Testing and Materials; 1991. pp. 355-364 - 23.
Gong P, Wilke BM, Fleischmann S. Soil-based phytotoxicity of 2,4,6-trinitrotoluene to higher plants. Archives of Environmental Contamination and Toxicology. 1999; 36 :152-157 - 24.
Dane F, Dalgıç Ö. The effects of fungicide benomyl (Benlate) on growth and mitosis in onion ( Allium cepa L.) root apical meristem. Acta Biologica Hungarica. 2005;56 :119-128 - 25.
Aksoy Ö, Erbulucu T, Vatan E. Effects of wastewater from olive oil and milk industry on growth and mitosis in Allium cepa root apical meristem. Journal of Applied Biological Sciences. 2011;5 (3):75-78 - 26.
Aksoy Ö, Deveci A. The investigation of the cytotoxic effects of some pesticides on soybean ( Glycine max L.). Cytologia. 2012;77 (4):475-483 - 27.
Tütünoğlu B, Aksoy Ö. Morphological and anatomical effects of the herbicide roundup on Zea mays (Poaceae). Journal of Functional and Environmental Botany. 2014;4 (2):71-76 - 28.
Zackrisson O, Nilsson MC. Allelopathic effects by Empetrum hermaphroditum on seed germination of two boreal tree species. Canadian Journal of Forest Research. 1992;22 (9):1310-1319. DOI: 10.1139/x92-174 - 29.
An J, Zhou Q, Sun Y, Xu Z. Ecotoxicological effects of typical personal care products on seed germination and seedling development of wheat ( Triticum aestivum L.). Chemosphere. 2009;76 :1428-1434 - 30.
Kapanen A, Itavaara M. Ecotoxicity tests for compost applications. Ecotoxicology and Environmental Safety. 2001; 49 :1-16 - 31.
Aksoy Ö, Erbulucu T, Öner S, Tekeli İB. Phytotoxic And genotoxic effects of water samples taken from the eastern channel of Kocaeli on Vicia faba andZea mays . Fresenius Environmental Bulletin. 2012;21 (7):1820-1826 - 32.
Aksoy Ö. The phytotoxic and genotoxic effects of olive oil and milk industry waste water on Elodea canadensis . Journal of Functional and Environmental Botany. 2013;3 (1): 43-53 - 33.
Helfrich P, Chefetz B, Hadar Y, Chen Y, Schnabl H. A novel method for determining phytotoxicity in compost. Compost Science and Utilization. 1998; 6 :6-13 - 34.
Gundersson CA, Kostuk JM, Mitcell HG, Napolitano GE, Wicker LF, Richmond JE, Stewart AJ. Multispecies toxicity assessment of compost produced in bioremediation of an explosives-contaminated sediment. Environmental Toxicology and Chemistry. 1997; 16 :2529-2537 - 35.
Maynard AA. Utilization of MSW compost in nursery stock production. Compost Science and Utilization. 1998; 6 :38-44 - 36.
Maluszynska J, Juchimiuk J. Plant genotoxicity: A molecular cytogenetic approach in plant bioassays. Arhiv za Higijenu Rada i Toksikologiju. 2005; 56 :177-184 - 37.
Eastmond DA. Sister chromatid exchanges. In: Wexler P. editor. Encyclopedia of Toxicology. 2nd ed. San Diego: Academic Press. 2005. pp. 19-20 - 38.
Iqbal M. Vicia faba bioassay for environmental toxicity monitoring: A review. Chemo-sphere. 2016;144 :785-802 - 39.
Feretti D, Zerbini I, Zani C, Ceretti E, Moretti M, Monarca S. Allium cepa chromosome aberration and micronucleus tests applied to study genotoxicity of extracts from pesticide-treated vegetables and grapes. Food Additives and Contaminants. 2007;24 (6):561-572 - 40.
Stich HF, Lam P, Lo LW, Koropatnick DJ and San HC. The search for relevant short-term bioassays for chemical carcinogens: The tribulation of a modern sisyphus. Canadian Journal of Genetics and Cytology. 1975; 17 :471-492 - 41.
Levan A. The effect of colchicine on root mitoses in Allium. Hereditas. 1938; 24 :471-486 - 42.
De Souza CP, Guedes TA, Fontanetti CS. Evaluation of herbicides action on plant bioindicators by genetic biomarkers: a review. Environmental Monitoring and Assessment. 2016; 188 (12):694. DOI: 10.1007/s10661-016-5702-8 - 43.
Ma TH, Grant WF. The Tradescantias—adventurous plants. The Herbarist. 1982; 48 :36-44 - 44.
Rank J, Nielsen MH. Evaluation of the Allium anaphase–telophase test in relation to genotoxicity screening of industrial wastewater. Mutation Research. 1994; 312 :17-24 - 45.
Cotelle S, Masfaraud JF, Férard JF. Assessment of the genotoxicity of contaminated soil with the Allium/Vicia-micronucleus and the Tradescantia-micronucleus assays. Mutation Research. 1999; 426 :167-171 - 46.
Chen G, White PA. The mutagenic hazards of aquatic sediments: A review. Mutation Research. 2004; 567 :151-225 - 47.
De Simone C, Tomati U, Galli E, Owczarek M, De Marco A, D’Ambrosio C, Alianello F, Cortellini L. Evaluation of toxic and genotoxic activity of some composts from different origin. Fresenius Environmental Bulletin. 2009; 9 :683-690 - 48.
Miao CM, Fu R, Yang D, Zheng L. Vicia root micronucleus assay on the clastogenicity of water samples from the Xiaoqing River in Shandong Province of the People's Republic of China. Mutation Research. 1999; 426 :143-145 - 49.
Monarca S, Zani C, Richardson SD, Thruston AD, Moretti JR, Feretti MD, Villarini MA. New approach to evaluating the toxicity and genotoxicity of disinfected drinking water. Water Reserach. 2004; 38 :3809-3819 - 50.
Khadra A, Pinelli E, Lacroix MZ, Bousquet-Melou A, Hamdi H, Merlina G, Guiresse M, Hafidi M. Assessment of the genotoxicity of quinolone and fluoroquinolones contaminated soil with the Vicia faba micronucleus test. Ecotoxicology and Environmental Safety. 2012;76 (2):187 - 51.
Ma TH. The role of plant systems for the detection of environmental mutagens and carcinogens. Mutation Research. 1999; 437 :97-100 - 52.
Rodrigues GS. Bioensaios de toxicidade genética com Tradescantia. Jaguariúna: Embrapa Meio Ambiente; 1999 - 53.
Gichner T, Veleminský J. Monitoring the genotoxicity of soil extracts from two heavily polluted sites in Prague using Tradescantia stamen hair and micronucleus (MNC) assays. Mutation Research. 1999; 426 :163-166 - 54.
Cebulska–Wasilewska A, Plewa MJ. Short term bioassay based on gene mutations level in Tradescantia cells (Trad-SH). In: Maluszynska J, Plewa M, editors. Bioassays in plant cells for improvement of ecosystem and human health. Katowice: Wydawnictwo Uniwersytetu Śląskiego. 2003. pp: 31-38 - 55.
Ma TH, Cabrera GL, Gill BS, Sandhu SS, Vandenberg AL, Salamone MF. Tradescantia micronucleus bioassay. Mutation Research. 1994; 310 :221-230 - 56.
Wilson DM, Thompson LH. Molecular mechanisms of sister-chromatid exchange. Mutation Research. 2007; 616 :11-23 DOI: 10.1016/j.mrfmmm.2006.11.017 - 57.
Cortès F, Andersson HC. Analysis of SCEs in Vicia faba chromosomes by a simple fluorescent plus Giemsa technique. Hereditas. 1987;107 :7-13 - 58.
Cortès F, Escalza P, Mateos S, Diaz-Recasens M. Factors affecting the production of SCEs by maleic hydrazide in root-tip chromosomes of Allium cepa . Mutation Research. 1987;192 :125-130 - 59.
Dimitrov B. Relationship between sister-chromatid exchanges and heterochromatin or DNA replication in chromosomes of Crepis capillaris. Mutation Research. 1987; 190 :271-276 - 60.
Maluszynska JB. Chromosomes of Crepis capillaris (L.) Waller. in vivo and in vitro. Katowice: Prace Naukowe Uniwersytetu Śląskiego nr 147; 1990 - 61.
Kolano B, Wolny E, Maluszynska J. Fluorescent in situ hybridisation in plant mutagenesis. In: Maluszynska J, Plewa M, editors. Bioassays in plant cells for improvement of ecosystem and human health. Katowice: Wydawnictwo Uniwersytetu Śląskiego; 2003. pp: 139-148 - 62.
Weiss H, Maluszynska J. Chromosomal rearrangement in autotetraploid plants of Arabidopsis thaliana . Hereditas. 2000;133 :255-261 - 63.
Maluszynska J, Juchimiuk J, Wolny E. Chromosomal aberrations in Crepis capillaris cell selected by FISH. Folia Histochem et Cytobiologia. 2003;41 :101-104 - 64.
Havel L, Durzan DJ. Apoptosis in plants. Botanica Acta: Journal of the German Botanical Society. 1996; 109 :268-277 - 65.
Ansari B, Coates PJ, Greenstein BD, Hall PA. In situ end labeling detects DNA strand breaks in apoptosis and other physiological and pathological states. Journal of Pathology. 1993; 170 :1-8 - 66.
Kanoh M, Takemura G, Misao J, Hayakawa Y, Aoyama T, Nishigaki K, Noda T, Fujiwara T, Fukuda K, Minatoguchi S, Fujiwara H. Significance of myocytes with positive DNA in situ nick end-labeling (TUNEL) in hearts with dilated cardiomyopathy. Not apoptosis but DNA repair. Circulation. 1999; 99 :2757-2764 - 67.
Juchimiuk J, Maluszynska J. Detection of DNA fragmentation caused by chemical mutagens using the TUNEL test. In: Maluszynska J, Plewa M, editors. Bioassays in Plant Cells for Improvement of Ecosystem and Human Health. Katowice: Wydawnictwo Uniwersytetu Śląskiego; 2003. pp. 133-138 - 68.
Jaloszynski P, Kujawski M, Szyfter K. Elektroforeza pojedynczych komórek (comet assay) – użyteczna technika badania uszkodzeń DNA. Postępy Biologii Komórki. 1996; 23 :339-354 - 69.
Gichner T. Comet assay in higher plants. In: Maluszynska J, Plewa M, editors. Bioassays in plant cells for improvement of ecosystem and human health. Katowice: Wydawnictwo Uniwersytetu Śląskiego; 2003. pp. 123-132 - 70.
Navarrete MH, Carrera P, Miguel M, Torre C. A fast comet assay variant for solid tissue cells. The assessment of DNA damage in higher plants. Mutation Research/Genetic Toxicology and Environmental Mutagenesis. 1997; 389 :271-277 - 71.
Pourrut B, Pinelli E, Celiz Mendiola V, Silvestre J, Douay F. Recommendations for increasing alkaline comet assay reliability in plants. Mutagenesis. 2015; 30 (1):37-43. DOI: 10.1093/mutage/geu075 - 72.
Agarwal M, Shrivastava N, Padh H. Advances in molecular marker techniques and their applications in plant sciences. Plant Cell Reports. 2008; 27 :617-631 - 73.
Aksoy Ö, Erbulucu T, Özen F, Deveci A. Genetic variation in critically endangered plant Amsonia orientalis Decne. Journal of Biodiversity and Environmental Sciences. 2013;3 (1):44-53 - 74.
Cenkci S, Yıldız M, Ciğerci İH, Konuk M, Bozdağ A. Toxic chemicals-induced genotoxicity detected by random amplified polymorphic DNA (RAPD) in bean ( Phaseolus vulgaris L.) seedlings. Chemosphere. 2009;76 :900-906 - 75.
Liu W, Yang YS, Zhou Q, Xie L, Li P, Sun T. Impact assessment of cadmium contamination on rice ( Oryza sativa L.) seedlings at molecular and population levels using multiple biomarkers. Chemosphere. 2007;67 :1155-1163 - 76.
Liu Z, Hossain GS, Islas-Osuna MA, Mitchell DL, Mount DW. Repair of UV damage in plants by nucleotide excision repair: Arabidopsis UVH1 DNA repair gene is a homolog of Saccharomyces cerevisiae Rad1. The Plant Journal. 2000;21 :519-528. DOI: 10.1046/j.1365-313x.2000.00707 - 77.
Mediouni C, Houlné G, Chaboute ME, Ghorbel MH, Jemal F. Cadmium and copper genotoxicity in plants. In: Biosaline Agriculture and High Salinity Tolerance. Basel: Birkhäuser; 2008. pp. 325-333. DOI: 10.1007/978-3-7643-8554-5-30 - 78.
Hu X, Jiang M, Zhang J, Zhang A, Lin F, Tan M. Calcium–calmodulin is required for abscisic acid-induced antioxidant defense and functions both upstream and downstream of H2O2 production in leaves of maize ( Zea mays ) plants. New Phytologist. 2007;173 :27-38. DOI: 10.1111/j.1469-8137.2006.01888 - 79.
Pfosser M, Heberle-Bors E, Amon A and Lelley T. Evaluation of sensitivity of flow cytometry in detecting aneuploidy in wheat using disomic and ditelosomic wheat–rye addition lines. Cytometry. 1995; 21 :387-393. DOI:10.1002/cyto.990210412 - 80.
Lane Rayburn A, Wetzel JB. Flow cytometric analyses of intraplant nuclear DNA content variation induced by sticky chromosomes. Cytometry. 2002; 49 (1):36-41. DOI:10.1002/cyto.10135 - 81.
Monteirono MS, Rodriguez E, Loureiro J, Mann RM, Soares AMVM & Santos C. Flow cytometric assessment of Cd genotoxicity in three plants with different metal accumulation and detoxification capacities. Ecotoxicology and Environmental Safety. 2010; 73 :1231-1237