Cloning and high expression of full length hTRF1 in E. Coli

Abstract

Objective: To express human telomeric repeat binding factor (TRF1) at high level in E. coli. Method: Two primers were designed with Kpn I and EcoR I sites respectively, TRF1 cDNA fragments was amplified and cloned into plasmid pET29α, each step was confirmed by sequencing and restriction endonuclease map analysis. And the recombinant plasmid pET29α-TRFl was then transformed into E. coli BL21 (DE3) PlysS. Fusion protein was purified by S-protein Kit and checked by SDS-PAGE and by western blot. Result: E. coli BL21 (DE3) PlysS expressing high level of 30 KD partial TRF1 was obtained, and TRF1 fusion protein was purified. The optimal induction time was at 2.5 h. Excessive expression system was established and 18.6% inductive protein was obtained. Conclusion: The expressed protein can be used for producing both polyclonal and monoclonal antibodies and for further study of the function and structure of TRF1 and its association with malignant tumor and leukemia.