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United States Department of Agriculture

Agricultural Research Service

Research Project: PHYSIOLOGICAL MECHANISMS THAT DETERMINE CROP RESPONSE TO IRRIGATION, DISEASE AND PRODUCTION PRACTICES Title: Modified method for combined DNA and RNA isolation from peanut and other oil seeds

Authors
item Dang, Phat
item Chen, Charles

Submitted to: Molecular Biology Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 9, 2012
Publication Date: February 1, 2013
Repository URL: http://DOI 10.1007/s11033-012-2204-9
Citation: Dang, P.M., Chen, C.Y. 2013. Modified method for combined DNA and RNA isolation from peanut and other oil seeds. Molecular Biology Reports. 40:1563-1568.

Interpretive Summary: peanuts are an important oilseed crop worldwide, containing about 50% oil of dry seed weight. Germinating or developing seeds are often challenged with diseases or adverse environmental conditions that could ultimately influence yield and quality. RNA isolation from plant seeds is a prerequisite for many seed specific gene expression studies and DNA is necessary in marker-assisted selection and other genetic studies. We describe a modified method to isolate both RNA and DNA from the same seed tissue and have been successful with several oil seeds including peanut, soybean, sunflower, canola, and oil radish. Quality of isolated nucleic acids was determined based on spectrophotometric measurement as well as gel-electrophoresis analysis. This research will facilitate the understanding of molecular signals produced by the seed during normal development and under disease or adverse environmental conditions; and the new information can lead to the development of new peanut varieties with enhanced seed agronomic characteristics.

Technical Abstract: Isolation of good quality RNA and DNA from seeds is difficult due to high levels of polysaccharides, polyphenols, and lipids that can degrade or co-precipitate with nucleic acids. Standard RNA extraction methods utilizing guanidinium-phenol-chloroform extraction has not shown to be successful. RNA isolation from plant seeds is a prerequisite for many seed specific gene expression studies and DNA is necessary in marker-assisted selection and other genetic studies. We describe a modified method to isolate both RNA and DNA from the same seed tissue and have been successful with several oil seeds including peanut, soybean, sunflower, canola, and oil radish. An additional LiCl precipitation step was added to isolate both RNA from DNA from the same seed tissues. High quality was observed based on A260/A280 and A260/A230 ratios above 2.0 and distinct bands on gel-electrophoresis. RNA was shown to be suitable for RT-PCR based on Actin or 60S ribosomal primer amplification and DNA was shown to have a single band on gel electrophoresis analysis. This result shows that RNA and DNA isolated using this method can be appropriate for molecular studies in peanut and other oil containing seeds.

Last Modified: 11/26/2014
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