Preparation and antigenicity analysis of 38 ku protein of mycobacterium tuberculosis / 中国实用内科杂志

ABSTRACT

Objective To Clone,express,purify 38 ku protein of mycobacterium tuberculosis,to study its immunological characteristics,and to evaluate its potenial value for serodiagnosis of tuberculosis.Methods Extract DNA from standard strain of H37Rv as the template,amplify gene of 38 ku protein by PCR,insert to PET-30a(+)and construct the recombinant plasmid,express 38 ku protein in E.coli BL21(DE3),purify by Nickel affinity chromatography,at last get target proteins of higher purity,analyze its immol/Lunological characteristics by Western blotting and ELISA technology from Feb.2003 to Mar.2004. Results The clone was analyzed at the nucleotide lever and showed the same DNA sequence coding for natural 38 ku protein. The recombinant protein expressed in inclusion body in E.coli BL21(DE3). The purity of terget protein was 92.7% by Nickel affinity chromatography,Western blotting assays showed that the recombinant protein had satisfactory antigenicity . 38 ku protein detected TB postive and negative refference serum based on the mechanism of indirect ELISA,results showed that the sensitivity was 80.5%(33/41)and the specificity reached to 96%(25/26).Conclusion The recombinant protein expressed in inclusion body in E.coli BL21(DE3)and had satisfactory antigenicity,and might be selected as one of serodiagnostic antigen of tuberculosis.