CTAB Method to Isolate DNA from Silica-Dried Frond Tissues of Several Tree Fern Species from Peninsular Malaysia
Keywords:
DNA isolation, CTAB, Silica-Gel Drying, Genomic DNA, PteridophytesAbstract
Extraction from field specimens of high-quality DNA often involves processing under liquid nitrogen that sometimes is not readily available in some laboratories. In this paper, a method of extracting DNA from silica-dried frond tissues of several tree fern species is described as existing protocols for genomic DNA isolation have their restrictions due to the existence of high content of tacky polysaccharides and polyphenols. Such protocols contaminate the DNA extracted with a viscous matrix, preventing for further use especially PCR performance. Polysaccharides and polyphenols were extracted in our adapted CTAB method before DNA precipitation and isopropanol was used to precipitate the genomic DNA. It was then amplified for testing with the non-coding primer trnL intron and trnL-F intergenic spacer. The highest concentration of DNA was from 0.03 g of frond tissues stored at -20 °C (205.80 ng/μL) for 24 hours, and even the lowest concentration obtained by this procedure was capable of generating enough PCR product for sequencing. The results showed the efficiency and reliability of the modified method as opposed to the unmodified method, suggesting that the combination of silica drying increases the quality and quantity of the DNA extracted.
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References
Dempster, E.L., et al., Rapid DNA extraction from ferns for PCR–based analyses. Biotechniques, 1999. 27(1): p. 66-68.DOI: https://doi.org/10.2144/99271bm13.
Saidin, S., A.A. Bakar, and B.M.M. Zain, Prevalence and associated risk factors of Entamoeba histolytica, E. dispar and E. moshkovskii infection among Orang Asli communities in Slim River, Perak. Journal of Science and Mathematics Letters, 2020. 8(2): p. 22-35.
Male, A.S., F. Kato, and C.M. Mukankusi, DNA extraction from silica gel preserved common bean (Phaseolus vulgaris L.) leaves. Genetics and Molecular Research, 17(4). 2018.DOI: https://doi.org/10.5897/AJB2018.16620.
Chase, M.W. and H.H. Hills, Silica gel: an ideal material for field preservation of leaf samples for DNA studies. Taxon, 1991. 40(2): p. 215-220.DOI: https://doi.org/10.2307/1222975.
Doyle, J.J., Isolation of plant DNA from fresh tissue. Focus, 1990. 12: p. 13-15.
Liston, A., et al., A method for collecting dried plant specimens for DNA and isozyme analyses, and the results of a field test in Xinjiang, China. Annals of the Missouri Botanical Garden, 1990. 77(4): p. 859-863.DOI: https://doi.org/10.2307/2399681.
Rogers, S.O. and A.J. Bendich, Extraction of DNA from milligram amounts of fresh, herbarium and mummified plant tissues. Plant molecular biology, 1985. 5(2): p. 69-76.DOI: https://doi.org/10.1007/BF00020088.
Narzary, D., et al., A rapid and effective method for isolation of genomic DNA from small amount of silica-dried leaf tissues. National Academy Science Letters, 2015. 38(5): p. 441-444.DOI: https://doi.org/10.1007/s40009-015-0357-5.
Ppg, I., A community‐derived classification for extant lycophytes and ferns. Journal of systematics and evolution, 2016. 54(6): p. 563-603.DOI: https://doi.org/10.1111/jse.12229.
Taberlet, P., et al., Universal primers for amplification of three non-coding regions of chloroplast DNA. Plant molecular biology, 1991. 17(5): p. 1105-1109.DOI: https://doi.org/10.1007/BF00037152.
Yen, K.H., et al., Detection of Porcine Based Materials in Processed Food Using Polymerase Chain Reaction Method. Journal of Science and Mathematics Letters, 2018. 6: p. 61-66.DOI: https://doi.org/10.37134/jsml.vol6.6.2018.
Adibah, A.B., W.L. Ng, and S.G. Tan, The Malay Peninsula as a barrier to gene flow in an Asian horseshoe crab species, Carcinoscorpius rotundicauda Latreille. Biochemical Systematics and Ecology, 60, 204-210. 2015.DOI: https://doi.org/10.1016/j.bse.2015.04.026.
Doyle, J.J. and J.L. Doyle, A rapid DNA isolation procedure for small quantities of fresh leaf tissue, 19, 11–15. 1987.
Korall, P., et al., A molecular phylogeny of scaly tree ferns (Cyatheaceae). American Journal of Botany, 2007. 94(5): p. 873-886.DOI: https://doi.org/10.3732/ajb.94.5.873.
Li, F.-W., et al., rbcL and matK earn two thumbs up as the core DNA barcode for ferns. PLoS One, 2011. 6(10): p. e26597.DOI: https://doi.org/10.1371/journal.pone.0026597.
Oldfield, S., International trade in tree ferns and evaluation on the application of CITES. World Conservation Monitoring Centre, Cambridge, UK, 1995.
Rout, S.D., T. Panda, and N. Mishra, Ethnomedicinal studies on some pteridophytes of similipal biosphere reserve, Orissa, India. International Journal of Medicine and Medical Sciences, 2009. 1(5): p. 192-197.
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