[Prokaryotic expression and purification of SPAG4L, a novel human testis gene].

Objective To express SPAG4L,a novel human testis gene in E.coli and purify it's fusion protein.Methods The fragment encoding SPAG4L126-379 was amplified by RT-PCR and the PCR products were cloned into PUCm-T vectors.After digestion by EcoR I and Hind III,the fragment was subcloned into PQE-30,a prokaryotic expression vector with 6×His tag.The recombinant plasmid PQE-30-SPAG4L was sequenced and transformed into E.coli M15.The expression of his-tagged fusion protein was induced by IPTG.The fusion protein was identified by Western blotting and purified using Ni-NTA magnetic agarose beads.Results The recombinant plasmid PQE-30-SPAG4L was constructed successfully and expressed in E.coli M15.The fusion protein SPAG4Lwith 6×his-tag was confirmed by Western blotting.The micro-scale purification system of 6×His-tagged SPAG4Lprotein was established and purified fusion protein was obtained.Conclusion The recombinant plasmid PQE-30-SPAG4L can be expressed in vitro and used for studying the biological function of SPAG4L in spermatogenesis.