Isolation of high-quality RNA from plant seeds
DOI:
https://doi.org/10.21638/spbu03.2021.205Abstract
The apple (Malus domestica Borkh.) is one of the major fruit tree crops, but it hasn’t been well-studied as a breeding object for molecular investigations. It is important to develop reliable and rapid methods that allow the preparation of plant material for future research. We introduce a quick and simple method for isolating high-quality RNA from lipid-rich apple seeds (M. domestica cv. Golden Delicious). Our method does not employ highly toxic reagents, because we exclude phenol, 2-mercaptoethanol and others. The chemical composition of the extraction buffer is simple and has a minimum level of toxicity. We showed that, in chaotropic conditions (i.e., with lithium chloride-urea), silica (SiO2) can bind with the lipids and RNA will remain in the solution. The extracted RNA was of high quality and we successfully used it for synthesizing cDNA and RT-PCR. The protocol developed by us can be useful for researchers working with RNA extraction from plant seeds.
Keywords:
Malus domestica, seeds, RNA, lipids, silica, RT-PCR
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References
Antonova, O. S., Korneva, N. A., Belov, Yu.V., and Kurochkin, V. E. 2010. Methods of nucleic acids purification and separation in molecular biology (review). Nauchnoe priborostroenie 20(1):3–9. (In Russian)
Asif, M., Trivedi, P., Solomos, T., and Tucker, M. 2006. Isolation of high-quality RNA from apple (Malus domestica) fruit. Journal of Agricultural and Food Chemistry 54(15):5227–5229. https://doi.org/10.1021/jf053137n
Birtic, S. and Kranner, I. 2006. Isolation of high-quality RNA from polyphenol-polysaccharide- and lipid-rich seeds. Phytochemical Analysis 17:144–148. https://doi.org/10.1002/pca.903
Codex Alimentarius (FAO/WHO). 2009. Codex standard for named vegetable oils codex stan 210-1999. http://www.codexalimentarius.org
Dal Cin, V., Danesin, M., Rizzini, F. M., and Ramina, A. 2005. RNA extraction from plant tissues. Molecular Biotechnology 31:113–119. https://doi.org/10.1385/MB:31:2:113
Ferrero, S., Carretero-Paulet, L., Mendes, M. A., Botton, A., Eccher, G., Masiero, S., and Colombo, L. 2015. Transcriptomic signatures in seeds of apple (Malus domestica L. Borkh) during fruitlet abscission. PLoS ONE 10(3):e0120503. https://doi.org/10.1371/journal.pone.0120503
Fromm, M., Bayha, S., Carle, R., and Kammerer, D. R. 2012. Characterization and quantitation of low and high molecular weight phenolic compounds in apple seeds. Journal of Agricultural and Food Chemistry 60(5):1232–1242. https://doi.org/10.1021/jf204623d
Gambino, G., Perrone, I., and Gribaudo, I. 2008. A rapid and effective method for RNA extraction from different tissues of grapevine and other woody plants. Phytochemical Analysis 19:520–525. https://doi.org/10.1002/pca.1078
Gasic, K., Hernandez, A., and Korban, S. S. 2004. RNA extraction from different apple tissues rich in polyphenols and polysaccharides for cDNA library construction. Plant Molecular Biology Reporter 22:437–438. https://doi.org/10.1007/BF02772687
Goulao, L. F. and Oliveira, C. M. 2007. Molecular identification of novel differentially expressed mRNAs up-regulated during ripening of apples. Plant Science 172: 306–318. https://doi:10.1016/j.plantsci.2006.09.006
Huang, C., Picimbon, J. F., Li, L. H. Q., Li, Z., Liu, Q., and Liu, W. 2012. An efficient method for total RNA extraction from peanut seeds. Russian Journal of Plant Physiology 59(1):143–147. https://doi.org/10.1134/S1021443712010074
Islam, M. R. and Banu, S. 2019. An improved cost-effective method of RNA extraction from Aquilaria malaccensis. Acta Scientific Agriculture 3(2):30–38.
Korte, K. and Casey, L. 1982. Phospholipid and neutral lipid separation by one-dimensional thin-layer chromatography. Journal of Chromatography B: Biomedical Sciences and Applications 232:47–53. https://doi.org/10.1016/S0378-4347(00)86006-5
Lan, T., Yao, B., Shen, Y., Wang, X. 2013. Isolation of high-quality total RNA from lipid-rich seeds. Analytical Biochemistry 438:11–13. https://doi.org/10.1016/j.ab.2013.03.012
Lay-Yee, M., DellaPenna, D., and Ross, G. S. 1990. Changes in mRNA and protein during ripening in apple fruit (Malus domestica Borkh. cv Golden Delicious). Plant Physiology 94:850–853. https://doi.org/10.1104/pp.94.2.850
Livak, K. J. and Schmittgen, T. D. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCt method. Methods 25:402–408. https://doi.org/10.1006/meth.2001.1262
Lu, Y. and Yeap Foo, L. 1998.Constitution of some chemical components of apple seed. Food Chemistry 61(1–2):29–33. https://doi.org/10.1016/S0308-8146(97)00123-4
Masek, T., Vopalensky, V., Suchomelova, P., and Pospisek, M. 2005. Denaturing RNA electrophoresis in TAE agarose gels. Analytical Biochemistry 336:46–50. https://doi.org/10.1016/j.ab.2004.09.010
Moser, C., Gatto, P., Moser, M., Pindo, M., and Velasco, R. 2004. Isolation of functional RNA from small amounts of different grape and apple tissues. Molecular Biotechnology 26:95–99. https://doi.org/10.1385/MB:26:2:95
Paula, M. F. B., Ságio, S. A., Lazzari, F., Barreto, H. G., Paiva, L.V., and Chalfun-Junior, A. 2012. Efficiency of RNA extraction protocols in different types of coffee plant tissues. Coffee Science 7(3):284–293.
Plotnikov, V. K. and Bakaldina, N. B. 1996. Differential stability of zein mRNA in developing corn kernel. Plant Molecular Biology 31:507–515. https://doi.org/10.1007/BF00042224
Portillo, M., Fenoll, C., and Escobar, C. 2006. Evaluation of different RNA extraction methods for small quantities of plant tissue: combined effects of reagent type and homogenization procedure on RNA quality-integrity and yield. Physiologia Plantarum 128:1–7. https://doi.org/10.1111/j.1399-3054.2006.00716.x
Prudnikov, S. M., Vityuk, B. Ya., and Zverev, L.V. 2003. Application of the method of nuclear magnetic relaxation to determine the moisture and oil content of agricultural materials. Pishchevaya promyshlennost' 2:42–44. (In Russian)
Radenkovs, V., Kviesis, J., Juhnevica-Radenkova, K., Valdovska, A., Püssa, T., Klavins, M., and Drudze, I. 2018. Valorization of wild apple (Malus spp.) by-products as a source of essential fatty acids, tocopherols and phytosterols with antimicrobial activity. Plants 7(90):1–18. https://doi.org/10.3390/plants7040090
Rott, M. E. and Jelkmann, W. 2001. Characterization and detection of several filamentous viruses of cherry: adaptation of an alternative cloning method (DOP-PCR), and modification of an RNA extraction protocol. European Journal of Plant Pathology 107:411–420. https://doi.org/10.1023/A:1011264400482
Saito, T., Wang, S., Ohkawa, K., Ohara, H., Ikeura, H., Ogawa, Y., and Kondo, S. 2017. Lipid droplet-associated gene expression and chromatin remodelling in LIPASE 5′-upstream region from beginning-to mid-endodormant bud in ‘Fuji’ apple. Plant Molecular Biology 95:441–449. https://doi.org/10.1007/s11103-017-0662-0
Šķipars, V., Šņepste, I., Krivmane, B., Veinberga, I., and Ruņģis, D. 2014. A method for isolation of high-quality total RNA from small amounts of woody tissue of Scots pine. Baltic Forestry 20(2):230–237.
Sundyreva, M. A., Stepanov, I. V., Suprun, I. I., and Ushakova, Ya. V. 2018. A modified protocol of RNA isolation from mature leaves of grapes for RT-PCR. Scientific Journal of KubSAU 143(09):1–15. https://doi.org/10.21515/1990-4665-143-012
Tai, H. H., Pelletier, C., and Beardmore, T. 2004. Total RNA isolation from Picea mariana dry seed. Plant Molecular Biology Reporter 22:93. https://doi.org/10.1007/BF02773357
Tian, H.-L., Zhan, P., and Li, K.-X. 2010. Analysis of components and study on antioxidant and antimicrobial activities of oil in apple seeds. International Journal of Food Sciences and Nutrition 61(4):395–403. https://doi.org/10.3109/09637480903535772
Tong, Z., Qu, S., Zhang, J., Wang, F., Tao, J., Gao, Z., and Zhang, Z. 2012. A modified protocol for RNA extraction from different peach tissues suitable for gene isolation and real-time PCR analysis. Molecular Biotechnology 50(3):229–236. https://doi.org/10.1007/s12033-011-9433-3
Xu, M., Zhang, B., Yao, H. S., and Huang, M. R. 2009. Isolation of high quality RNA and molecular manipulations with various tissues of Populus. Russian Journal of Plant Physiology 56(5):716–719. https://doi.org/10.1134/S1021443709050197 (In Russian)
Xu, Y., Fan, M., Ran, J., Zhang, T., Sun, H., Dong, M., Zhang, Z., and Zheng, H. 2016. Variation in phenolic compounds and antioxidant activity in apple seeds of seven cultivars. Saudi Journal of Biological Sciences 23:379–388. https://doi.org/10.1016/j.sjbs.2015.04.002
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Published
2021-06-30
How to Cite
Mishko, A., Sundyreva, M., Stepanov, I., Efimenko, S., Plotnikov, V., & Nenko, N. (2021). Isolation of high-quality RNA from plant seeds. Biological Communications, 66(2), 144–150. https://doi.org/10.21638/spbu03.2021.205
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Articles of Biological Communications are open access distributed under the terms of the License Agreement with Saint Petersburg State University, which permits to the authors unrestricted distribution and self-archiving free of charge.
