A rapid TRIzol-based two-step method for DNA-free RNA extraction from Arabidopsis siliques and dry seeds.
Extraction of high-quality RNA from Arabidopsis seeds has been a challenge. Here we report a two-step TRIzol-based procedure for RNA extraction from Arabidopsis siliques and dry seeds. This procedure employs a modified, high pH (pH 9.5) extraction buffer. High pH plus the addition of either DTT or beta-mercaptoethanol in the extraction buffer effectively inhibits RNase activity during the extraction, and removes most polysaccharides, polyphenols and other insoluble material. TRIzol reagent was subsequently used to purify the RNA. Using this procedure we isolated high-quality DNA-free RNA samples without DNase I treatment from Arabidopsis seeds or siliques in less than 3 h.
Meng
Ling
L
Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720-3102, USA. mengling@berkeley.edu
Feldman
Lewis
L
D017360:Arabidopsis / Q000737:chemistry* / Q000235:genetics; D005591:Chemical Fractionation / Q000379:methods*; D018744:DNA, Plant / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D005638:Fruit / Q000737:chemistry* / Q000235:genetics; D006146:Guanidines / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008623:Mercaptoethanol / Q000737:chemistry; D010636:Phenols / Q000737:chemistry*; D018749:RNA, Plant / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012639:Seeds / Q000737:chemistry* / Q000235:genetics
D018744:DNA, Plant; D006146:Guanidines; D010636:Phenols; D018749:RNA, Plant; C411644:trizol; D008623:Mercaptoethanol; D004229:Dithiothreitol