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).
Lamarcq
Laurence H
LH
Clontech Laboratories, Mountain View, CA 94043, USA.
Scherer
Bradley J
BJ
Phelan
Michael L
ML
Kalnine
Nikolai N
NN
Nguyen
Yen H
YH
Kabakova
Tatyana
T
Chen
Xiaoyi
X
Tan
Marcia
M
Chang
Cynthia
C
Berlon
Charina
C
Campos-Gonzalez
Roberto
R
Gao
Guo-Jian
GJ
Golz
Stefan
S
Vysotski
Eugene S
ES
Farmer
Andrew A
AA
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