[Construction of pcDNA3-HERG-G572R expression vector and establishment of a cell line stably expressing HKE-HERG-G572R].

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.