Establishment and identification of HUVEC cell strains with over-expression and low expression of RACK 1
- VernacularTitle:RACK1过表达和低表达HUVEC细胞系的建立和鉴定
- Author:
Li ZHANG
;
Xiongfei JIA
;
Hua NIU
;
Yue FENG
;
Yuzhu SONG
;
Wanglong ZHANG
;
Shengying LI
;
Mingjiang WU
;
Xiaoqin MAO
- Publication Type:Journal Article
- Keywords:
RACK1;
over-expression;
RNA interference;
HUVEC strain
- From:
Basic & Clinical Medicine
2015;(2):218-223
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish several human umbilical vein endothelial cell ( HUVEC ) strains with over-ex-pression or low expression of receptor for activated C kinase 1 ( RACK1 ) , which will provide an effective tool for future studying the function of RACK1 in arrhythmia.Methods The full-length cDNA sequence of RACK1 gene was amplified and inserted into pIRES2-EGFP.At the same time, designed and synthesised complementary DNA sequences of 3 pairs of short hairpin structure and a pair of negative control sequence , then subcloned into the plas-mid pGenesil-1 .The HUVEC cells were transfected with these plasmids and screened by using G 418 .And the expression of RACK1 mRNA and protein in the cells were assayed by qRT-PCR and Western blot , respectively . Results RACK1 eukaryotic expression vector and siRNA expression vectors of RACK 1 were constructed success-fully.After a 48 h transfection of HUVEC cells with the recombinant vectors and G 418 selection, the positive cell clones were obtained .qRT-PCR and Western blot showed that over-expression vector and interference vectors could effectively enhanced and knocked-down RACK1 expression in HUVEC strains .Conclusions HUVEC cell strains with over-expression and low expression of RACK 1 have been successfully established .
