1.Construction of pcDNA3-HERG-G572R expression vector and establishment of a cell line stably expressing HKE-HERG-G572R.
Yang YANG ; Na HUANG ; Ling GAO ; Su'e CHANG ; Bo GUO ; Lili HU ; Tusheng SONG ; Chen HUANG
Journal of Southern Medical University 2014;34(3):308-311
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.
Base Sequence ; Ether-A-Go-Go Potassium Channels ; genetics ; Gene Expression ; Genetic Vectors ; HEK293 Cells ; metabolism ; Humans ; Mutation ; Transfection
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