title: Isolation of high quality RNA and construction of a suppression subtractive hybridization library from ramie (Boehmeria nivea L. Gaud.).
abstract: Isolation of high quality RNA from ramie (Boehmeria nivea L. Gaud.) is difficult due to its high levels of polyphenols, polysaccharides, pectin, fat, wax and other secondary metabolites. A modified procedure based on guanidinium isothiocyanate for RNA preparation of ramie was developed in this study. High concentrations (5%, v/v) of guanidinium isothiocyanate, PVP-4000, sodium citrate and sodium lauryl sarcosinate and beta-mercaptoethanol were used in the extraction buffer, together with a low pH sodium acetate (pH 4.0) added to improve the RNA quality. The average yield was about 400 microg RNAg(-1) fresh leaves. One SSH library which was induced by ramie anthracnose was constructed by utilizing the RNA extracted through the present method. These results showed that our protocol was applicable for RNA isolation from recalcitrant ramie tissues.
authors.affiliation: College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, 430070, Hubei Province, Poeple's Republic of China.
authors.forename: Xu-xia
authors.initials: XX
authors.lastname: Wang
authors.forename: Bo
authors.initials: B
authors.lastname: Wang
authors.forename: Li-jun
authors.initials: LJ
authors.lastname: Liu
authors.forename: Xiao-ping
authors.initials: XP
authors.lastname: Cui
authors.forename: Jin-yu
authors.initials: JY
authors.lastname: Yang
authors.forename: Hui
authors.initials: H
authors.lastname: Wang
authors.forename: Hui
authors.initials: H
authors.lastname: Jiang
authors.forename: Bei-bei
authors.initials: BB
authors.lastname: Luo
authors.forename: Zhu
authors.initials: Z
authors.lastname: Long
authors.forename: Wen-xiang
authors.initials: WX
authors.lastname: Dou
authors.forename: Na
authors.initials: N
authors.lastname: Zhang
authors.forename: Ding-xiang
authors.initials: DX
authors.lastname: Peng
mesh_terms: D015152:Blotting, Northern; D032402:Boehmeria / Q000235:genetics* / Q000382:microbiology; D002999:Clone Cells; D020231:Colletotrichum / Q000502:physiology; D004587:Electrophoresis, Agar Gel; D015723:Gene Library; D009693:Nucleic Acid Hybridization / Q000379:methods*; D018749:RNA, Plant / Q000302:isolation & purification* / Q000592:standards*; D013053:Spectrophotometry
chemical_list: D018749:RNA, Plant
references.citation: Biotechnol Lett. 2005 May;27(9):629-33
references.pmid: 15977069
references.citation: Proc Natl Acad Sci U S A. 1996 Jun 11;93(12):6025-30
references.pmid: 8650213
references.citation: Plant Physiol. 2005 Aug;138(4):2165-73
references.pmid: 16024691
references.citation: Mol Genet Genomics. 2005 Jun;273(4):326-35
references.pmid: 15812649
references.citation: Biotechnol Lett. 2006 Apr;28(8):587-91
references.pmid: 16614897
references.citation: Biotechniques. 1992 Jul;13(1):52-4, 56
references.pmid: 1503775
references.citation: Anal Biochem. 2008 Mar 15;374(2):426-8
references.pmid: 18166144
references.citation: Biotechnol Lett. 2005 Dec;27(23-24):1841-5
references.pmid: 16328977
references.citation: Nat Protoc. 2006;1(2):581-5
references.pmid: 17406285
references.citation: Dokl Biol Sci. 2008 Nov-Dec;423:440-2
references.pmid: 19213431
references.citation: Anal Biochem. 1994 Nov 15;223(1):7-12
references.pmid: 7535022