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[Gene cloning and prokaryotic expression of the PID domain of human DOC-2 molecule]
Shu-Juan Liu1, Wei-Guo Zheng, Zong-Ling Ji
1Department of Obstetrics and Gynecology,Xijing Hospital, Fourth Military Medical University, Xi'an 710032 China.
Aim:
To clone the PID domain of human DOC-2 (nDOC-2) and express it in E.coli DH5alpha.
Methods:
The cDNA fragment encoding the PID domain of nDOC-2 was amplified by RT-PCR from normal human ovarian tissue and cloned to pUC19. The DNA fragment from the pUC19-nDOC-2 digested with BamH I and EcoR I was ligated to the BamH I/EcoR I digested prokaryotic expression vector pGEX-4T-1.The expression of fusion protein was induced with IPTG and the expressed product was identified by SDS-PAGE.
Results:
(1)The sequencing and endonucleases digestion analysis showed that the fragment of nDOC-2 gene was insert into vectors pUC19 and pGEX-4T-1;(2)SDS-PAGE showed the nDOC-2 gene had been expressed in E.coli DH5alpha.
Conclusion:
The PID domain of nDOC-2 was expressed successfully in prokaryote, which makes preparation for further researching the function of DOC-2 and preparing antibodies to DOC-2 protein.
