Cloning,expression and immunity analysis of transketolase of Echinococcus granulosus / 中国血吸虫病防治杂志

ABSTRACT

Objective To obtain the prokaryotic expression of transketolase genes and analyze its value as a diagnostic anti-gen for echinococcosis.Methods TK gene was amplified by PCR and cloned into prokaryotic vector pMD19-EgTK,and then subcloned into the expression vector pET-28a.The target gene TK prokaryotic expression plasmid pET-28a was constructed and transferred into BL21. The purified protein was identified by SDS-PAGE and Western blotting. The blood samples of patients with cystic echinococcosis(CE group),alveolar echinococcosis(AE group)and healthy people(healthy group)were collected and detected by ELISA with the recombinant EgTK protein as a diagnostic antigen.Results The recombinant plasmid pET-28a (+)-EgTK was constructed successfully,and there was a band around 70 kDa by using Western blotting.ELISA showed that the difference among the 3 groups of sera reaction A450was significantly different(F=44.47,P<0.01),and the A450values of the CE group(1.46±0.41)and AE group(1.28±0.29)were higher than that of the healthy group(0.66 ± 0.23),but there was no significant difference between the former two.Conclusion The recombinant EgTK protein is better to distinguish the echinococ-cosis group and healthy group,but it can't do a differential diagnosis between CE and AE cases.