Construction of pcDNA3-HERG-G572R expression vector and establishment of a cell line stably expressing HKE-HERG-G572R.
- Author:
Yang YANG
1
;
Na HUANG
;
Ling GAO
;
Su'e CHANG
;
Bo GUO
;
Lili HU
;
Tusheng SONG
;
Chen HUANG
Author Information
- Publication Type:Journal Article
- MeSH: Base Sequence; Ether-A-Go-Go Potassium Channels; genetics; Gene Expression; Genetic Vectors; HEK293 Cells; metabolism; Humans; Mutation; Transfection
- From: Journal of Southern Medical University 2014;34(3):308-311
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct the pcDNA3-HERG-G572R expression vector and establish a cell line stably expressing HKE-HERG-G572R.
METHODSHERG-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.
RESULTSThe 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).
CONCLUSIONThe protocol can be used to construct the cell line stably expressing HKE-HERG-G572R to provide a cell model for studying individualized therapy.
