[Cloning, expression and purification of Schistosoma japonicum ribosomal protein S4 as well as the preliminary study of the diagnostic value of the recombinant protein].
OBJECTIVE
To express and purify Schistosoma japonicum ribosomal protein S4(SjRPS4) in Escherichia coli, and assess its value in immunodiagnosis of Schistosomiasis japonica.


METHODS
Gene fragment of SjRPS4 was amplified by screening the cercaria cDNA library of Schistosoma japonicum. The target gene was cloned into the expressive vector pQE30 and transformed into E. coli M15. The recombinant protein expression was induced by isopropylthio-β-D-galactoside (IPTG). This fusion protein was purified by Ni(2+)-NTA chromatography and identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blot and ELISA.


RESULTS
The plasmid pQE30/SjRPS4 was constructed successfully and expressed a SjRPS4 fusion protein in E. coli as showing a single special band on SDS-PAGE gel at Mr 30 × 10(3) position. It reached a purity of above 90% after purification. The Western blot result confirmed that the recombinant protein could specifically react with the serum samples from patients of schistosomiasis. Detecting the serum of Schistosomiasis japonica patients by ELISA, the sensitivity and specificity of the ELISA method were 90.91% (70/77) and 92.59% (25/27), the positive rate of recombinant protein expression was 67.30% (70/104). There was no cross-reaction with paragonimiasis patients' serum.


CONCLUSIONS
Protein SjRPS4 was successfully cloned and expressed, and it was confirmed that SjRPS4 antibodies were valuable in the diagnosis of Schistosomiasis japonica.
The Key Laboratory of Immune and Control Schistosomiasis, Xiangya School of Medicine, Central South University, Changsha 410078, China.
Dong-mei
DM
Gao
Shi-ping
SP
Wang
Lu-xin
LX
Yu
Zhuo
Z
He
Xiu-chun
XC
Chen
D000595:Amino Acid Sequence; D000818:Animals; D000909:Antibodies, Helminth / Q000097:blood; D000947:Antigens, Helminth; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics; D012269:Ribosomal Proteins / Q000235:genetics*; D012549:Schistosoma japonicum / Q000235:genetics*; D012554:Schistosomiasis japonica / Q000175:diagnosis* / Q000235:genetics; D012680:Sensitivity and Specificity
D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; C031798:ribosomal protein S4