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Simple and Efficient Protocol for RNA and DNA extraction from Rice (Oryza sativa L.) for Downstream Applications

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Author Affiliations

  • 1Centre for Advanced Studies in Plant Biotechnology and Genetic Engineering, Department of Biosciences, Saurashtra University, Rajkot, Gujarat,

Int. Res. J. Biological Sci., Volume 4, Issue (2), Pages 62-67, February,10 (2015)

Abstract

Plant DNA and RNA extraction is difficult due to the presence of metabolites that interfere with isolation procedures and downstream applications. In the present study DNA and RNA extraction protocol standardized from rice (Oryza sativa L.). Isolation of DNA using Triton-X-100 based extraction method with PVPP treatment efficiently removes metabolites and yield with high quality DNA from rice coleoptiles.RNA isolation methods based on TRIzol was suitable for rice leaf whereas in seed, TrisHCl (pH 9.0) with mercaptoethanol protocol gave best results. The isolated DNA and RNA proved to be suitable for PCR and RT-PCR amplification, respectively. The technique is reproducible and can be applied for PCR based markers identification and gene expression studies.

References

  1. Fang G.H. and Grumet R., A quick and inexpensive method for removing polyccharides from plant genomic DNA, Biotechniques.,13(1), 52-56 (1992)
  2. Doyle J. J. and Doyle J. L., Isolation of plant DNA from fresh tissue, Focus., 12, 13–15 (1990)
  3. Pal. S. Jain.S. Saini. N. and Jain R.K., DNA isolation from milled rice samples for PCR-based molecular marker analysis, Rice Genetics Newsletters., 18, 94-97 (2001)
  4. Chen Wen-yue., Cui Hai-rui., Bao Jin-song., Zhou Xiang-sheng and Shu Qing-yao., A Simplified Rice DNA Extraction Protocol for PCR Analysis, Rice Science., 13(1), 67-70, (2006)
  5. Rodrigues S.M., Virg΄nia L.F., Soares Tahise, de Oliveira M., Abelmon S. Gesteira, Wagner C., Otoni and Marcio G.C., Isolation and Purification of RNA from tissues rich in polyphenols, polysaccharides, and pigments of Annatto (Bixa orellana L.), Mol Biotechnol., 37, 220–224 (2007)
  6. Gambino G, Perrone I and Gribaudo I., A rapid and effective method for RNA extraction from different tissues of grapevine and other woody plants, Phytochem Anal., 19, 520–525 (2008)
  7. Xu J, Aileni M, Abbagani S and Zhang P., A reliable and efficient method for total RNA isolation from various members of spurge family (Euphorbiaceae), Phytochem Anal., 210, 395–8 (2010)
  8. Guifeng Wang, Gang Wang, Xiaowei Zhang, Fang Wang and Rentao Song., Isolation of high quality RNA from Cereal seeds containing high levels of Starch, Phytochem. Anal., 23, 159–163 (2012)
  9. Mandaliya V.B., Pandya R.V. and Thaker V.S., Comparison of cotton DNA extraction methods for the high yield and quality from various cotton tissues, J. Cotton Res. Dev., 24 (1), 9-12 (2010)
  10. Sghaier zidani., Genomic DNA extraction method from pearl millet (Pennisetum glaucum) leaves, African Journal of Biotechnology., 4(8), 862-866 (2005)
  11. Wilkie S.E., Issac P.G. and Slater R.J., Random amplification polymorphic DNA (RAPD) markers for genetic analysis in Allium, Theor. Appl. Genet., 86, 497-504 (1993)
  12. Murray M.G and Thomson W.F., Rapid isolation of high molecular weight plant DNA, Nucl Acids Res., 19, 4321–4325 (1980)
  13. Paterson A.H., Brubaker C.L. and Wendel J.F., A rapid method for extraction of cotton (Gossypium spp.) genomic DNA suitable for RFLP or PCR analysis, Plant Mol. Biol. Rep., 11(2), 122-127,(1993)
  14. Sangha J.S., Gu K., Kaur J. and Yin Z., S An improved method for RNA isolation and cDNA library construction from immature seeds of Jatropha curcas L, BMC Res Notes,3(1), 126 (2010)
  15. Chomczynski P. and Sacchi N., Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction, Anal. Biochem., 162, 156-9 (1987)
  16. Sambrook J. and Russell D.W., In Molecular cloning A Laboratory Manual Cold Spring Harbor Laboratory Press,(2001)
  17. Dellaporta S.L., Wood J. and Hicks J.B., A plant DNA mini preparation version II, Plant Mol Biol Rep., 19-21 (1983)
  18. Arif I.A., Bakir M.A., Khan. H.A., Farhan A.H., Homaidan A.A., Bahkali A.H.M., Sadoon Al and Shobrak M., Application of RAPD for molecular characterization of plant species of medicinal value from an arid environment, Genet. Mol. Res., 9(4), 2191-2198 (2010)
  19. Pikbart M.J. and Villeponteau B., Suppression of PCR amplification by high levels of RNA, Biotechniques., 14,24-25(1993)
  20. Niu C., Kebede H., Auld D.L., Woodward J.E., Burow G. and Wright R.J., A safe inexpensive method to isolate high quality plant and fungal DNA in an open laboratory environment, Afr. J. Biotech.,7(16), 2818-2822 (2008)
  21. De V.S., Hoge H and Bisseling T., Isolation of total and polysomal RNA from plant tissues, In Plant Molecular Biology Manual, B6, Gelvin SB, Schilperoort RA, Verma DPS (eds)., Kluwer Academic Publishers., 13(1993)