Extraction of high quality of RNA and construction of a suppression subtractive hybridization (SSH) library from chestnut rose (Rosa roxburghii Tratt).
Chestnut rose (Rosa roxburghii Tratt) is a rare fruit crop of promising economical importance in fruit and ornamental exploitation in China. Isolation of high quality RNA from chestnut rose is difficult due to its high levels of polyphenols, polysaccharides and other compounds, but a modified CTAB extraction procedure without phenol gave satisfactory results. High concentrations of PVP (2%, w/v), CTAB (2%, w/v) and beta-mercaptoethanol (4%, v/v) were used in the extraction buffer to improve RNA quality. The average yield was about 200 microg RNA g(-1) fresh leaves. The isolated RNA was of sufficient quality for construction of suppression subtraction hybridization (SSH) library, which allowed the isolation of several pathogen-induced defense genes.
Xu
Qiang
Q
National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070 Wuhan, PR China.
Wen
Xiaopeng
X
Tao
Nengguo
N
Hu
Zhiyong
Z
Yue
Hailin
H
Deng
Xiuxin
X
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D015723:Gene Library; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics / Q000302:isolation & purification*; D027826:Rosa / Q000235:genetics*; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D012333:RNA, Messenger; D018749:RNA, Plant