Recombination and identification of human alpha B-crystallin  

作　　者：Rui Wang Ze-Hua Chen Yi Wang Hou-Bin Huang Si-Jun Fan Lan-Lan Chen 

机构地区：[1]Department of Ophthalmology,Hainan Branch of PLA General Hospital,Sanya 572000,Hainan Province,China [2]Chongqing Aier General Hospital,Aier School of Ophthalmology,Central South University,Chongqing 400020, China [3]PLA Rocket Force General Hospital,Beijing 100088,China

出　　处：《International Journal of Ophthalmology(English edition)》2018年第12期1916-1921,共6页国际眼科杂志（英文版）

基　　金：Supported by National Natural Science Foundation of China Grant (No.81270996);Science and Technology Project Foundation of Hainan Province (No.ZDYF201631);Health Science and Technology Innovation Project Foundation of Sanya (No.2016YW22)

摘　　要：AIM: To recombine the human alpha B-crystallin(αBcrystallin) using gene cloning technology and prokaryotic expression vector and confirm the biological activity of recombinant human αB-crystallin. METHODS: Cloning the human αB-crystallin cDNA according to the nucleotide sequence of the human αBcrystallin, constructing the pET-28/CRYAB prokaryotic expression plasmid by restriction enzyme digestion method, and stably expressing transformed into the Escherichia coli(E. coli) DH5 alpha. The recombinant human αB-crystallin was purified by Q sepharose. By enzyme digestion analysis, Western blotting and sequencing, the recombinant human αB-crystallin was identified and the activity of its molecular protein was detected.RESULTS: Compared with the gene bank(GeneBank), the cloned human sequence of human αB-crystallin cDNA has the same open reading frame. Identification and sequencing of the cloned human αB-crystallin cDNA in prokaryotic expression vector confirmed the full length sequence, and the vector was constructed successfully. The E. coli containing plasmid pET-28/CRYAB induced by isopropyl-β-D-thiogalactoside successfully expressed the human αB-crystallin. Insulin confirmed that the recombinant human αB-crystallin has a molecular chaperone activity. CONCLUSION: The prokaryotic expression vector pET-28/CRYAB of recombinant human αB-crystallin issuccessfully constructed, and the recombinant human αBcrystallin with molecular chaperone activity is obtained, which lay a foundation for the research and application of the recombinant human αB-crystallin and its chaperone activity.AIM: To recombine the human alpha B-crystallin(αBcrystallin) using gene cloning technology and prokaryotic expression vector and confirm the biological activity of recombinant human αB-crystallin. METHODS: Cloning the human αB-crystallin cDNA according to the nucleotide sequence of the human αBcrystallin, constructing the pET-28/CRYAB prokaryotic expression plasmid by restriction enzyme digestion method, and stably expressing transformed into the Escherichia coli(E. coli) DH5 alpha. The recombinant human αB-crystallin was purified by Q sepharose. By enzyme digestion analysis, Western blotting and sequencing, the recombinant human αB-crystallin was identified and the activity of its molecular protein was detected.RESULTS: Compared with the gene bank(GeneBank), the cloned human sequence of human αB-crystallin cDNA has the same open reading frame. Identification and sequencing of the cloned human αB-crystallin cDNA in prokaryotic expression vector confirmed the full length sequence, and the vector was constructed successfully. The E. coli containing plasmid pET-28/CRYAB induced by isopropyl-β-D-thiogalactoside successfully expressed the human αB-crystallin. Insulin confirmed that the recombinant human αB-crystallin has a molecular chaperone activity. CONCLUSION: The prokaryotic expression vector pET-28/CRYAB of recombinant human αB-crystallin issuccessfully constructed, and the recombinant human αBcrystallin with molecular chaperone activity is obtained, which lay a foundation for the research and application of the recombinant human αB-crystallin and its chaperone activity.

关 键 词：human alpha B-crystallin GENE vector construction IDENTIFICATION recombination 

分 类 号：R[医药卫生]