title: Modified method for combined DNA and RNA isolation from peanut and other oil seeds.
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 and DNA from the same seed tissues. High quality nucleic acids were observed based on A(260)/A(280) and A(260)/A(230) ratios above 2.0 and distinct bands on gel-electrophoresis. RNA was shown to be suitable for reverse transcriptase polymerase chain reaction 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.
authors.affiliation: National Peanut Research Laboratory, USDA-ARS, PO Box 509, 1011 Forrester Dr. SE, Dawson, GA 39842, USA. Phat.Dang@ars.usda.gov
authors.forename: Phat M
authors.initials: PM
authors.lastname: Dang
authors.forename: Charles Y
authors.initials: CY
authors.lastname: Chen
mesh_terms: D010367:Arachis / Q000737:chemistry / Q000235:genetics*; D029688:Brassica napus / Q000737:chemistry / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics / Q000302:isolation & purification*; D044943:Fatty Acid Desaturases / Q000235:genetics; D018628:Gene Dosage; D005838:Genotype; D006368:Helianthus / Q000737:chemistry / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics; D018749:RNA, Plant / Q000235:genetics / Q000302:isolation & purification*; D031224:Raphanus / Q000737:chemistry / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012639:Seeds / Q000737:chemistry / Q000235:genetics*; D013025:Glycine max / Q000737:chemistry / Q000235:genetics
chemical_list: D018744:DNA, Plant; D010940:Plant Proteins; D018749:RNA, Plant; D044943:Fatty Acid Desaturases; C040448:delta-12 fatty acid desaturase