[Construction and identification of different stem shRNA expression vectors].
We constructed shRNA vectors with different stem length, and tested the silencing effectiveness in mouse cells and embryos. We designed interfering RNAs with stems of 21 bp, 27 bp, and 29 bp. The enhanced green fluorescent protein gene was used as target gene. The synthesized single strands were annealed and cloned into psiSTRIKE and the recombinant plasmids (EGFP-21 siRNA, EGFP-27 siRNA, and EGFP-29 siRNA) were transfected into the mouse embryonic fibroblast with lipofection. The mRNA expression level of the enhanced green fluorescent protein gene was checked by real-time quantitative PCR. The silencing effectiveness of the 29 bp shRNA vector was stronger than which of the 21 bp and 27 bp. The findings in this study are of interest for selecting the hairpins for mouse individuals.
Liu
Zhonghua
Z
Shaanxi Stem Cell Engineering and Technology Research Center, Shaanxi Key Laboratory of Molecular Biology for Agriculture, College of Veterinary Medicine, Northwest A & F University, Yangling 712100, China.
Qiao
Xianfeng
X
Xiao
Hongwei
H
Liu
Ximei
X
Wang
Huayan
H
Zheng
Xinmin
X
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D020868:Gene Silencing; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D018547:Plant Stems / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection
D034741:RNA, Small Interfering; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins