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KMID : 0380420140380040075
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Journal of Prventive Veterinary Medicine
2014 Volume.38 No. 4 p.75 ~ p.78
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Molecular cloning, expression and purification of Brucella abortus 544 phosphoglycerate kinase in a pMAL vector
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Reyes Alisha Wehdnesday Bernardo
Simborio Hannah Leah Tadeja
Hop Huynh Tan
Arayan Lauren Togonon
Min Won-Gi
Lee Hu-Jang
Chang Hong-Hee
Kim Suk
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Abstract
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Brucellosis is an important and re-emerging zoonotic disease worldwide. The prevention of human infection isachieved predominantly through the control of brucellosis in agricultural animals, which in turn depends on accurate diagnosisand vaccination. However, conventional serological diagnosis of brucellosis has several limitations, and currently availablevaccines for animals have several drawbacks, including the ability to cause infection in humans. Phosphoglycerate kinase(Pgk) is one of the specific proteins reactive with mouse sera in the early stage of Brucella infection, and deletion of thepgk gene in B. abortus strain 2308 resulted in extreme attenuation of this strain in vitro and in vivo. Furthermore, the B. abortus pgk mutant has been used as a live vaccine, and in challenge experiments, it induced protection that was superiorto that conferred by commercial strains. In this study, the pgk gene from Brucella abortus 544 was successfully amplified andcloned into a maltose binding protein fusion protein expression vector (pMAL). The recombinant protein was expressed inEscherichia coli DH5α and purified. The immunogenicity of purified recombinant B. abortus 544 Pgk (rPgk) was evaluatedby western blot analysis using Brucella-positive mouse sera. rPgk could be used as an antigenic component for futureserological tests and potential vaccine development.
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KEYWORD
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Brucella abortus, pgk, pMAL, cloning, expression
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