Cloning and Expression of Gene Encoding Extracellular Loop 2 of Tetraspanins of Schistosoma japonicum

Objective To clone and express the gene encoding extracellular loop 2(EC-2) of tetraspanins(TSPs) of Schistosoma japonicum(Chinese strain).Method An unknown ortholog of S.mansoni TSP-2 was screened from the ESTs of S.japonicum,and the corresponding gene of which was finally called Sj-tsp-2 after modifications.The interest gene synthesized was then directionally cloned into prokaryotic expression vector pET32a.The recombinant plasmid pET-32a-Sj-tsp-2 was transformed into competent E.coli BL21(DE3)and expressed in the presence of IPTG.Fusion protein was purified from E.coli lysates under nondenaturing conditions.Result DNA of recombinant plasmid pET-32a-Sj-tsp-2 was sequenced in its own right.SDS-PAGE analysis showed that the expressed fusion protein was around 26 kDa.More importantly,the results of Western blotting revealed that the interest protein Sj-tsp-2 could be specifically recognized by sera from mice vaccinated with recombinant protein Trx-Sj-tsp-2.Conclusion The successful expression and purification of recombinant protein Trx-Sj-tsp-2 make it possible for further research on animal protection experiments.