Cloning of the Gene Encoding Mycoplasma hyopneumoniae P36 Protein, Expression and Evaluating the Antigenicity of Recombinant Protein
Cloning of the Gene Encoding Mycoplasma hyopneumoniae P36 Protein, Expression and Evaluating the Antigenicity of Recombinant Protein
Nguyen Thi Thu Hien1, Dong Huu Rin2, Nguyen Xuan Hoa1, Le Viet Tuan Khanh2, Phung Thang Long1*, Dinh Thi Bich Lan1
ABSTRACT
This study aimed to clone the gene (432 bp) encoding M. hyopneumoniae P36 protein, express this protein in E. coli BL21(DE3), and evaluate its antigenicity as a base for further studies to against M. hyopneumoniae. The gene encoding M. hyopneumoniae P36 protein isolated from DNA of fresh lung tissue samples from PEP infected pigs was cloned into pGEM®-T Easy vector for sequencing, and then was inserted into pET28a vector to express the protein. The cloning result revealed that the gene encoding the M. hyopneumoniae P36 protein was 432 bp in length, and 99.07% similarity to the nucleotide sequence of P36 gene from strain M. hyopneumoniae 7448 recorded in GenBank (accession No. AE017244.1), corresponding to 144 amino acids. Expression of this protein in E. coli BL21 produced a fusion protein with molecular weight of approximately 20 kDa. Moreover, the antibodies against M. hyopneumoniae in sera taken from pigs that were immunized with the PEP inactivated vaccine (HYOGEN®) strong detected the recombinant M. hyopneumoniae P36 protein by ELISA, demonstrating that expressed recombinant P36 protein has the antigenicity and is specific for M. hyopneumoniae. In summary, we have successfully cloned the gene (432 bp) encoding the M. hyopneumoniae P36 protein. The expressed recombinant M. hyopneumoniae P36 protein in strain E. coli BL21 has the antigenicity and is specific for M. hyopneumoniae. Further experiments should be performed to evaluate the immunogenicity of M. hyopneumoniae P36 protein for use as an antigen to develop novel vaccines and bio-products (e.g., egg yolk antibody) against M. hyopneumoniae.
Keywords | Mycoplasma hyopneumoniae, porcine enzootic pneumonia, P36 protein, protein expression, recombinant protein, antigenicity
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November 2023
Vol. 11, Iss. 11, pp. 1757-1910
