Prokaryotic Expression and Identification of cTnI-linker-TnC Fusion Protein

doi:10.13523/j.cb.20150407

China Biotechnology

2015, Vol. 35

Issue (4): 48-53 DOI: 10.13523/j.cb.20150407

Prokaryotic Expression and Identification of cTnI-linker-TnC Fusion Protein

GONG Long-cai^1,2, LUO Zhen-ming^1,2, YANG Yan-qing^1,2, WANG Zhen-yu^1,2, XIANG Jun-jian^1,2, WANG Hong^1,2

1. Department of Bioengineering, Guangdong Province Key Laboratory of Molecular Immunology and Antibody Engineering, Jinan University, Guangzhou 510632, China;
2. Guangzhou Key Laboratory of Disease and Food Safety Rapid Test, Guangzhou 510632, China

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Abstract

The aim is to express and identify the fusion protein cTnI-linker-TnC, and detect its immunogenicity for preparation of monoclonal antibody of anti-cTnI-TnC. A short DNA sequence which codes 15 neutral amino acid residues was used to link cTnI and cTnC. Then, the recombinant prokaryotic expression vectors pET32a-cTnI-linker-TnC and pET28a-cTnI-linker-TnC were constructed to express the fusion protein cTnI-linker-TnC by transformed into E.coli BL21 cells respectively. And the fusion proteins cTnI-linker-TnC were identified by 12% SDS-PAGE and indirect ELISA test.At the same time, the concentrations of the inducing agent,expression times and temperatures were optimized to obtain high soluble target protein. At last, the commercial monoclonal antibodies of anti-cTnI and anti-cTnC were used to do the Western blot test and double antibody sandwich ELISA,and the serum of myocardial infarction patients were used to do the indirect ELISA. The results showed that the fusion protein cTnI-linker-TnC has a high soluble expression, and has reserved its immunogenicity well. This indicates the fusion protein cTnI-linker-TnC may replace the natural cTnI-TnC compound in the future.

Key words： cTnI-linker-TnC Fusion protein Prokaryotic expression Double antibody sandwich ELISA

Received: 30 December 2014 Published: 25 April 2015

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Cite this article:

GONG Long-cai, LUO Zhen-ming, YANG Yan-qing, WANG Zhen-yu, XIANG Jun-jian, WANG Hong. Prokaryotic Expression and Identification of cTnI-linker-TnC Fusion Protein. China Biotechnology, 2015, 35(4): 48-53.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20150407 OR https://manu60.magtech.com.cn/biotech/Y2015/V35/I4/48

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