[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 HKEHERG-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(P0.01).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.