Large-scale, high-throughput validation of short hairpin RNA sequences for RNA interference.
A method for high-throughput cloning and analysis of short hairpin RNAs (shRNAs) is described. Using this approach, 464 shRNAs against 116 different genes were screened for knockdown efficacy, enabling rapid identification of effective shRNAs against 74 genes. Statistical analysis of the effects of various criteria on the activity of the shRNAs confirmed that some of the rules thought to govern small interfering RNA (siRNA) activity also apply to shRNAs. These include moderate GC content, absence of internal hairpins, and asymmetric thermal stability. However, the authors did not find strong support for position specific rules. In addition, analysis of the data suggests that not all genes are equally susceptible to RNA interference (RNAi).
Clontech Laboratories, Mountain View, CA 94043, USA.
Laurence H
LH
Lamarcq
Bradley J
BJ
Scherer
Michael L
ML
Phelan
Nikolai N
NN
Kalnine
Yen H
YH
Nguyen
Tatyana
T
Kabakova
Xiaoyi
X
Chen
Marcia
M
Tan
Cynthia
C
Chang
Charina
C
Berlon
Roberto
R
Campos-Gonzalez
Guo-Jian
GJ
Gao
Stefan
S
Golz
Eugene S
ES
Vysotski
Andrew A
AA
Farmer
D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D006801:Humans; D016133:Polymerase Chain Reaction; D012313:RNA / Q000737:chemistry*; D034622:RNA Interference
D017931:DNA Primers; D012313:RNA