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Updated: Sep 20, 2026
Functional Cloning Using a Xenopus Oocyte Expression System
Published on: January 30, 2016
[Cloning and expression of specific antigen genes of Ancylostoma caninum]
Xiang-rong Guo1, Hai-chou Xue, Tie-hua Li
1Institute of Parasitic Disease, Chinese Center for Disease Control and Prevention, Shanghai 200025.
Objective:
To search for the gene encoding specific antigen of Ancylostoma caninum that induces host's protective immunity.
Methods:
A lambda ZAP II cDNA library of A. caninum was screened with sera from dogs immunized subcutaneously with hookworm larvae(L3). After sequencing, insert gene (AcAg) from positive clones was ligated into PUC18 and PET28C. Recombinant pET28C plasmid was induced to expressed protein in the E. coli BL21. The characteristic of recombinant protein is analyzed by SDS-PAGE and Western blotting assay.
Results:
Five positive clones were obtained, and proved to be the same. The insert gene (AcAg) in pET28C vector expressed a recombinant protein of about 43 kDa. Using Western blotting assay, this protein was recognized by the sera from dog immunized with Ancylostoma caninum third-stage infected larvae and used for screening library.
Conclusion:
The AcAg, which exhibits 35% homologous to Caenorhabditis elegans gene unc-89, is a novel specific antigen of A. caninum. Its ability to elicit the protective immunity and the probability of the recombinant protein as a vaccine need to be further evaluated.
