Construction of pcDNA3-HERG-G572R expression vector and establishment of a cell line stably expressing HKE-HERG-G572R
10.3969/j.issn.1673-4254.2014.03.04
- VernacularTitle:HERG基因G572R突变体表达载体的构建及其稳定转染细胞系的建立
- Author:
Yang YANG
1
;
Na HUANG
;
Ling GAO
;
Sue CHANG
;
Bo GUO
;
Lili HU
;
Tusheng SONG
;
Chen HUANG
Author Information
1. 西安交通大学医学院遗传学与分子生物学系
- Keywords:
HERG gene;
HEK293 cell;
transfection;
mutation
- From:
Journal of Southern Medical University
2014;(3):308-311
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the pcDNA3-HERG-G572R expression vector and establish a cell line stably expressing HKE-HERG-G572R. Methods HERG-G572R mutant fragment was constructed by over-lap extension PCR and validated by DNA sequencing. The HKE-HERG-G572R expression vector was constructed and transfected into HEK293 cells to obtain a cell line stably expressing HKE-HERG-G572R. Results The pcDNA3-HERG-G572R expression vector was successfully constructed and the cell line stably expressing HKE-HERG-G572R was established. Real-time PCR and Western blotting revealed a 632-fold HKE-HERG-G572R overexpression in the transfected HEK293 cells as compared with that in control HEK293 cells transfected with pcDNA3 (P<0.01).Conclusion The protocol can be used to construct the cell line stably expressing HKE-HERG-G572R to provide a cell model for studying individualized therapy.
