河流型水牛热休克蛋白70的原核表达及其对精液抗冻性的影响

doi:10.16431/j.cnki.1671-7236.2018.09.009

Abstract

Abstract:

The purpose of the research was to clone heat shock protein 70 (HSP70) gene CDS region,construct the HSP70 prokaryotic expression vector,and induce the expression of HSP70 fusion protein to further study the effect of HSP70 on the freezing semen motility of river buffalo.The buffalo sperm genome DNA was extracted as a template, the HSP70 gene CDS region was amplified by PCR,and the PCR products was connected with pET30a plasmid to construct recombinant prokaryotic expression plasmid and induce the expression of HSP70.The obtained HSP70 was added to the buffalo frozen semen and it's motility was evaluated.The results showed that the CDS region of HSP70 gene was amplified successfully with 2 155 bp in length.The SDS-PAGE and Western blotting analysis showed that the fusion protein had the molecular weight of 70 ku and could react with antibodies.The final concentration of 2,4 and 8 mg/mL HSP70 could improve the motilities of frozen semen,and 8 mg/mL HSP70 had the best effect on the motilities,but there were no significant differences among groups (P>0.05).In conclusion,the purified HSP70 obtained in this study had good quality,and could improve the frozen semen motility.The study laid a theoretical foundation for further studying the antifreeze protection mechanism of buffalo HSP70 protein.

Key words: river buffalo; HSP70; prokaryotic expression; sperm motility

CLC Number:

Q786

LI Fangfang, LONG Kaixu, FENG Xiangren, HUANG Xiangsheng, PAN Tangfeng, JIANG Mingsheng, WANG Guoli, DENG Zhuxin, JIANG Qinyang, GUO Yafen. Prokaryotic Expression of HSP70 and It's Effect on the Freezing Resistance of Semen in River Buffalo[J]. , 2018, 45(9): 2417-2424.

References

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Prokaryotic Expression of HSP70 and It's Effect on the Freezing Resistance of Semen in River Buffalo

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