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Issue:ISSN 2095-1353
           CN 11-6020/Q
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Your Position :Home->Past Journals Catalog->2019年56 No.2

Molecular cloning and prokaryotic expression of the odorant binding protein MpOBP8 in Microplitis pallidipes Szepligeti
Author of the article:YANG An1, 2** MEI Guo-Hong3 ZHANG Hao2 JIANG Jie-Xian2 JI Xiang-Yun2*** CHEN Yi-Juan2
Author's Workplace:1. College of Life Science and Fishery, Shanghai Ocean University, Shanghai 201306, China; 2. Eco-envirmental Protection Research Institute, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China;3. Jinshan District Agro-technology Extension Center, Shanghai 201500, China
Key Words:Microplitis pallidipes; MpOBP8; odorant binding protein; optimization expression
Abstract: 

[Objectives]  MpOBP8 is an odorant binding protein (OBP) of Microplitis pallidipes, which plays an essential role in the perception of external odors. The aim of this study was to clone the OBP, optimize its expression, and explore the molecular mechanism of olfactory recognition. [Methods]  Differential expression in different tissues was analyzed with semi-quantitative PCR. A fusion expression vector pET32a (+) -MpOBP8 was constructed using double enzyme digestion. The plasmid pET32a (+) -MpOBP8 was transferred into the Escherichia coli and the expression conditions were optimized. [Results]  The MpOBP8 gene (GenBank accession number: MG457156) was cloned. The open reading frame was 459 bp, which encoded a polypeptide of 152 amino acids. The protein molecular mass was predicted as 17.57 ku with an isoelectric point of 4.78. The expression of MpOBP8 was detected in the antennae with semi-quantitative PCR, but no expression was detected in the head, thorax or abdomen. The best induction temperature, induction time, and the final IPTG concentration were 25 , 6 h and 0.1 mmol·L-1, respectively, whereas the induction opportunity for protein expression at culture medium with OD600 was up to 0.1-0.3. We obtained most target protein under the above conditions and a high concentration by purification and concentration. [Conclusion]  We successfully cloned the MpOBP8 gene and optimized the conditions for its prokaryotic expression, obtaining a large amount of the odorant binding protein. These results provide a potential basis for exploring the molecular mechanism underlying olfactory recognition.

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