Development of an Efficient Protocol of RNA Isolation from Recalcitrant Tree Tissues
Tóm tắt
Isolation of RNA from recalcitrant tree tissues has been problematic due to large amounts of secondary metabolites and interfering compounds in their cells. We have developed an efficient RNA extraction method, which yielded high-quality RNA preparations from tissues of the lychee tree. The method reported here utilized EDTA, LSS, and CTAB to successfully inhibit RNase activities. It was found that a high ionic strength brought about by 2 M NaCl was necessary. In addition, secondary metabolites and other interfering compounds were effectively removed using sodium borate and PVPP under a deoxidized condition. The quality of purified RNA was tested by both RACE and Northern blotting analysis, ensuring that the RNA could be used for subsequent gene expression analysis. This method has been successfully applied to purify RNA from 15 other plant species. In conclusion, the protocol reported here is expected to have excellent applications for RNA isolation from recalcitrant plant tissues.
Tài liệu tham khảo
Ainsworth, C. (1994). Isolation of RNA from floral tissue of Rumex acetosa (Sorrel). Plant Molecular Biology Reporter, 12, 198–203.
Birtic, S., & Kranner, I. (2006). Isolation of high-quality RNA from polyphenol-, polysaccharide- and lipid-rich seeds. Phytochemical Analysis, 17, 144–148.
Chang, S., Puryear, J., & Caimey, J. (1993). A simple and efficient method for isolating RNA from pine trees. Plant Molecular Biology Reporter, 11, 113–116.
Chomczynski, P., & Sacchi, N. (1987). Single step method of RNA isolation by acid guanidinium thiocyanale-phenol-chloroform extraction. Analytical Biochemistry, 162, 156–159.
Chomczynski, P. (1993). A reagent for the single-step simultaneous isolation of RNA, DNA and proteins from cell and tissue samples. Biotechniques, 15, 532–537.
Claros, M. G., & Canovas, F. M. (1998). Rapid high quality RNA preparation from pine seedlings. Plant Molecular Biology Reporter, 16, 9–18.
Davy, A. J., Bishop, G. F., & Costa, C. B. (2001). Salicornia L. (Salicornia pusilla J. Woods, S. ramosissima J. Woods, S. europaea L., S. obscura P.W. Ball & Tutin, S. nitens P.W. Ball & Tutin, S. fragilis P.W. Ball & Tutin and S. dolichostachya Moss). Journal of Ecology, 89, 681–707.
Geuna, F., Hartings, H., & Scienza, A. (1998). A new method for rapid extraction of high quality RNA from recalcitrant tissues of grapevine. Plant Molecular Biology Reporter, 16, 61–67.
Ignacio, I. F., Leticia, P. E., Blondy, C. C., & Monica, R. C. (2006). Extraction of high-quality, melanin-free RNA from Mycosphaerella fijiensis for cDNA preparation. Molecular Biotechnology, 34, 45–50.
Ildiko, B., Janpeter, N., & Mlynarova, L. (1999). Isolation of high quality DNA and RNA from leaves of the carnivorous plant Drosera rotundifolia. Plant Molecular Biology Reporter, 17, 269–277.
Jelle, D. K., Isabel, R. R., Erik, V. B., Arne, H., & Denis, D. K. (2006). Efficient extraction of High-Quality total RNA from various hop tissues. Preparative Biochemistry and Biotechnology, 36, 355–362.
John, M. E. (1992). An efficient method for isolation of RNA and DNA from plants containing polyphenolics. Nucleic Acids Research, 20, 2381.
Kiefer, E., Werener, H., & Dieter, E. (2000). A simple and efficient protocol for isolation of functional RNA from plant tissues rich in secondary metabolites. Plant Molecular Biology Reporter, 18, 33–39.
Kim, S. H., & Hamada, T. (2005). Rapid and reliable method of extracting DNA and RNA from sweetpotato, Ipomoea batatas (L). Lam. Biotechnology Letter, 27, 1841–1845.
Li, J. H., Tang, G. H., Song, C. Y., Chen, M. J., Feng, Z. Y., & Pan, Y. J. (2006). A simple, rapid and effective method for total RNA extraction from Lentinula edodes. Biotechnology Letters, 28, 1193–1197.
Liu, J. J., Goh, C. J., Loh, C. S., Liu, P., & Pua, E. C. (1998). A method for isolation of total RNA from fruit tissues of banana. Plant Molecular Biology Reporter, 16, 1–6.
Logemann, J., Schell, J., & Willmitzer, L. (1987). Improved method for isolation of RNA from plant tissue. Analytical Biochemistry, 163, 16–20.
Malnoy, M., Reynoird, J. P., Mourgues, F., Cheverau, E., & Simoneau, P. (2001). A method for isolation total RNA from pear. Plant Molecular Biology Reporter, 19, 69–74.
Manickavelu, A., Kambara, K., Mishina, K., & Koba, T. (2007). An efficient method for purifying high quality RNA from wheat pistils. Colloid Surface, B54, 254–258.
Meisel, L., Fonseca, B., Gonzalez, S., Baezayates, R., Cambiazo, V., Campos, R., Gonzalez, M., Orelana, A., Retamales, J., & Silva, H. (2005). A rapid and efficient method for purifying high quality total RNA from peaches (prunus persica) for functional genomics analyses. Biological Research, 38, 83–88.
Portillo, M., Fenoll, C., & Escobar, C. (2006). Evaluation of different RNA extraction methods for small quantities of plant tissue. Physiologia Plantarum, 128, 1–7.
Riehl, T. E., & Ungar, I. A. (1982). Growth and ion accumulation in Salicornia europaea under saline field conditions. Oecologia, 54, 193–199.
Russell, B., Rathinasabapathi, B., & Hanson, A. D. (1998). Osmotic stress induces expression of choline monooxygenase in sugar beet and amaranth. Plant Physiology, 116, 859–865.
Salzman, R. A., Fujiya, T. K., Salzman, Z., Hasgawa, P. M., & Bressan, R. A. (1999). An improved RNA isolation method for plant tissues containing high levels of phenolic compounds or carbohydrates. Plant Molecular Biology Reporter, 17, 11–17.
Sambrook, J., Fritsch, E. F., & Maniatis, T. (1989). Molecular cloning: A laboratory manual (pp. 343–388). Cold Spring Harbour, New York: Cold Spring Harbour Laboratory Press.
Shi, H. Z., & Bressan, R. (2006). RNA extraction. Methods in Molecular Biology, 323, 345–348.
Su, X., & Gibor, A. (1988). A method for RNA isolation from marine macro-algae. Analytical Biochemistry, 174, 650–657.
Wan, C. Y., & Wilkins, T. A. (1994). A modified hot borate method significantly enhances the yield of high-quality RNA from cotton (Gossypium hirsutum L.). Analytical Biochemistry, 223, 7–12.
Xu, Q., Wen, X. P., Tao, N. G., Hu, Z. Y., Yue, H. L., & Deng, X. X. (2006). Extraction of high quality of RNA and construction of a suppression subtractive hybridization (SSH) library from chestnut rose (Rosa roxburghii Tratt). Biotechnology Letters, 28, 587–591.
Yamashita, Y., Sakurai, T., Kuno, N., Uchida, K., & Yokobayshi, T. (2005). RNA extraction method, RNA extraction reagent, and method for analyzing biological materials. United States Patent, 981521.
Zeng, Y., & Yang, T. (2002). RNA isolation from highly viscous samples rich in polyphenols and polysaccharides. Plant Molecular Biology Reporter, 20, 415–417.