[Cloning, expression and identification of endothelial differentiation-related factor-1 gene of Schistosoma japonicum].
OBJECTIVE
To clone and express endothelial differentiation-related factor (SjEDF)-1 gene of Schistosoma japonicum, analyze its immunogenicity and the stage-specific expression at different developmental stages of S. japonicum.


METHODS
Total RNA were extracted from eggs, cercariae, schistosomula and adult worms. The housekeeping gene SjActin was selected as the internal reference. According to the open reading frame for SjEDF-1 gene (GenBank accession number: AY336498), a pair of primers were designed to amplify the SjEDF-1 gene which was subcloned into pET-28a vector. The recombinant plasmid SjEDF-1/pET-28a was transformed into E. coli BL21 and induced with IPTG for expression. The recombinant protein was purified with Ni-NTA resin. The immune rabbit sera was prepared by immunizing New Zealand white rabbits with purified recombinant SjEDF-1 protein. Western blotting was used to analyze the immunogenicity and the expression level of SjEDF-1 at the different developmental stages.


RESULTS
The SjEDF-1 gene was detected with a band of 405 bp in eggs, schistosomula, female and male worms. The recombinant protein (rSjEDF-I) was expressed as inclusion bodies (M, 20 000). Western blotting analysis showed that the purified rSjEDF-1 protein was recognized by pooled sera of infected rabbits. The target protein was detected only in schistosomulum and adult worms.


CONCLUSIONS
The recombinant protein (rSjEDF-I) shows certain immunogenicity, and is detected only in schistosomula and adult worms.
Deng
Wang-Ping
WP
School of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Feng
Zheng
Z
Xu
Bin
B
Kong
Juan
J
Hu
Wei
W
D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D015801:Helminth Proteins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D010957:Plasmids; D011817:Rabbits; D012549:Schistosoma japonicum / Q000235:genetics* / Q000378:metabolism; D012554:Schistosomiasis japonica / Q000469:parasitology
D002148:Calmodulin-Binding Proteins; D015801:Helminth Proteins