Abstract | AIM: To induce the expression of FMDV receptor integrin beta6 subunit ligand-binding domain in E.coli and prepare the rabbit polyclonal antibody against it. METHODS: The fragment coding beta6 ligand-binding domain was amplified by PCR and doubly digested with BamH Iand Xho I. Then it was cloned into expression vector pGEX-4T-1 to obtain recombinant plasmid pGEX-4T-1-beta6LBD. After pGEX-4T-1-beta6LBD was transformed into E.coli BL21(DE3) and induced by IPTG, the expression of fusion proteins was identified by SDS-PAGE, with inclusion body prepared and fusion protein purified. Then new Zealand rabbits were immunized to prepare polyclonal antibody against beta6LBD. GST-beta6LBD antiserum was obtained and the specificity of polyclonal antibody was detected by Western blot. RESULTS: SDS-PAGE demonstrated that the fusion protein GST-beta6LBD was expressed with the expected molecular weight at 42 000. A single clear band of GST-beta6LBD fusion protein appeared in SDS-PAGE gel after purification. The titer of the polyclonal antibody was above 1:12 800 and it is of high specificity. CONCLUSION: The successful preparation of rabbit anti pig beta6LBD polyclonal antibody with high affinity and specificity will lay a foundation for further research into the function of integrin beta6 in FMDV infection.
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Authors | Shan-Dian Gao, Hui-Yun Chang, Jun-Zheng DU, Jing-Feng Wang, Jian-Hua Zhou, Jian-Yong Zhao, Qing-Ge Xie |
Journal | Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
(Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi)
Vol. 24
Issue 10
Pg. 975-8
(Oct 2008)
ISSN: 1007-8738 [Print] China |
PMID | 18845083
(Publication Type: English Abstract, Journal Article)
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Chemical References |
- Antibodies
- Integrin beta Chains
- Protein Subunits
- Receptors, Virus
- integrin beta6
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Topics |
- Animals
- Antibodies
(immunology, metabolism)
- Blotting, Western
- Electrophoresis, Polyacrylamide Gel
- Foot-and-Mouth Disease Virus
(metabolism, pathogenicity)
- Integrin beta Chains
(genetics, immunology, metabolism)
- Male
- Polymerase Chain Reaction
- Protein Subunits
(genetics, immunology, metabolism)
- Rabbits
- Receptors, Virus
(genetics, immunology, metabolism)
- Swine
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