Isolation of high quality RNA and construction of a suppression subtractive hybridization library from ramie (Boehmeria nivea L. Gaud.).
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.
Wang
Xu-xia
XX
College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, 430070, Hubei Province, Poeple's Republic of China.
Wang
Bo
B
Liu
Li-jun
LJ
Cui
Xiao-ping
XP
Yang
Jin-yu
JY
Wang
Hui
H
Jiang
Hui
H
Luo
Bei-bei
BB
Long
Zhu
Z
Dou
Wen-xiang
WX
Zhang
Na
N
Peng
Ding-xiang
DX
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
D018749:RNA, Plant
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