[Prokaryotic expression and identification of secretory proteins and peripheral membrane proteins of Schistosoma japonicum].

OBJECTIVE

To prokaryotic express and identify the recombinant plasmids containing the genes or gene segments which coding secreted or peripheral membrane protein of Schistosoma japonicum.

METHODS

A number of 28 pET32 (+) previous constructed recombinant plasmids containing these genes or segments were transferred into E. coli BL21 (DE3) strain for culture via the CaCl2 transformation method, and then induced by IPTG for prokaryotic expression. Then, the bacteria were treated by broking the wall with ultrasonic, and the bacterial suspension, supernatant and precipitation were detected with 12% SDS-PAGE. The size and distribution of these target proteins were determined. Finally, the expression of the recombinant fusion protein was further identified by the protein microarray combined with anti-His tag fluorescent antibody.

RESULTS

Under the inducing conditions of 0.1 mmol/L IPTG, 37 ℃, 200 r/min and 4 h, these fusion proteins were highly expressed in E. coli BL21 (DE3). The SDS-PAGE results showed all the 28 recombinant fusion proteins were expressed in different degrees, among which 2 were distributed in the supernatant, and the other 26 were distributed in precipitation, and the sizes of the fusion proteins were the same as that of prediction. The fluorescent signals of His antibody were successfully detected by confocal laser scanner.

CONCLUSIONS

A series of recombinant plasmids containing genes or gene segments that coding secreted protein and peripheral membrane protein of S. japonicum have been successfully expressed by using an efficient prokaryotic expression system of E. coli. It has established a foundation for the high throughput immunoscreening of the vaccine candidates or diagnostic antigens of S.japonicum.