Cloning of Bmi1 cDNA from mouse testis and its expression in E. coli BL21 / 中华男科学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To produce BMI1 polyclonal antibody, mouse Bmi1 cDNA was cloned from mouse testis and expressed in E. coli BL21.</p><p><b>METHODS</b>Bmi1 gene was amplified from mouse testis by RT-PCR and inserted into the prokaryotic expression vector pET-28c(+). Subsequently the recombined vector was transformed and expressed in E. coli BL21 (DE3) and the immunogenicity of recombined protein BMI1 (rBMI1) was tested by Western blot.</p><p><b>RESULTS</b>Mouse Bmi1 cDNA of 975 bp was successfully cloned and recombined. E. coli BL21 strains expressed rBMI1 were screened. The expression protein amounted to 12% of the total bacterial protein after induced with IPTG, which included inclusion body and soluble protein. Inclusion body was the major pattern of the expression that amounted to 71% of the insoluble protein. Western blot analysis showed that rBMI1 could be specially recognized by mouse monoclonal IgG1 anti-BMI1 and His-tag antibody.</p><p><b>CONCLUSION</b>There was expression of Bmi1 gene in mouse testis. Mouse Bmi1 cDNA was successfully cloned and expressed prokaryoticly.</p>