Prokaryotic expression and polyclonal antibody preparation of Hsc70 in Nile tilapia

doi:10.3969/j.issn.1004-1524.2019.08.07

›› 2019, Vol. 31 ›› Issue (8): 1272-1279.DOI: 10.3969/j.issn.1004-1524.2019.08.07

• Animal Science • Previous Articles Next Articles

Prokaryotic expression and polyclonal antibody preparation of Hsc70 in Nile tilapia

PAN Chuanyan, LIN Yong, FENG Pengfei, ZHANG Yongde, LUO Honglin^*

Guangxi Academy of Fishery Sciences, Guangxi Key Laboratory of Aquatic Genetic Breeding and Healthy Aquaculture, Nanning 530021, China

Received:2019-01-22 Online:2019-08-25 Published:2019-08-30

Abstract

Abstract: To express Hsc70 (heat shock cognate 70) of Nile tilapia via a prokaryotic expression system, the recombinant protein was purified and polyclonal antibody was prepared. Bioinformatics analysis of Hsc70 was analyzed, primers were designed, and then Hsc70 gene was amplified. A prokaryotic expression vector of Hsc70 was constructed and the Hsc70 recombinant protein was induced by IPTG.The recombinant protein was identified by SDS-PAGE and purified with a Ni-NTA. To obtain the polyclonal antibody, the purified Hsc70 recombinant protein was used to immunize Japanese white rabbits, the titer was determined by ELISA, and the specificity was analyzed by Western blot. The Hsc70 recombinant protein was mainly obtained from the inclusion body with a molecular weight of 75 ku. After the purified recombinant protein was immunized with Japanese white rabbits, the rabbit anti-Hsc70 polyclonal antibody was successfully obtained, whose titer was 1∶2 048 000 and the specificity was good. Hsc70 of Nile Tilapia expressed successfully in E. coli B21, and the recombinant protein and polyclonal antibody obtained in this study provided a molecular tool for further study the function of Hsc70.

Key words: Nile tilapia, Hsc70, prokaryotic expression, polyclonal antibody

CLC Number:

S917.4

PAN Chuanyan, LIN Yong, FENG Pengfei, ZHANG Yongde, LUO Honglin. Prokaryotic expression and polyclonal antibody preparation of Hsc70 in Nile tilapia[J]. , 2019, 31(8): 1272-1279.

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Prokaryotic expression and polyclonal antibody preparation of Hsc70 in Nile tilapia

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