Background: The aim of this research is to design and to clone the rhPTH gene in E. coli BL21 (DE3).
Methods: This research discloses a novel frame designed for efficient preparation of N-terminal 1-34 amino acids of hPTH as functional parts of this hormone. The frame constructed as an NcoI-BamH1 fragment encoding a His-tag and a chimeric fusion protein consisting of a fusion partner comprising of 52 amino acids belonging to Escherichia coli b- galactosidase (LacZ) gene, a cleavage site identified by Entropeptidase and rhPTH (1-34) gene fragment. Optimized frame was synthesized and ligated with pET28a vector under the control of T7 promoter, and then transformed in E. coli BL21 (DH5a) cells.
Results: Positive clones that released the mentioned frame by double digestion with NcoI-BamH1 enzymes were approved by sequencing.
Conclusion: We cloned designed gene in E. coli Bl21 (DH5a). We expect to express our designed gene to be very elevated after itsinduction by IPTG in Escherichia coli BL21 (DE3).