RESUMO
Objective: To obtain polycystin-1 intracellular region. Methods: cDNA of polycystin-1 intracellular region was generated by PCR and then cloned into pProEX Hta, which was prokaryotic expression vector. After verified by sequencing, the recombinant was transformed into E.coli host to express and purify the fusion protein by affinity chromatography. Results: 660 bp cDNA of polycystin-1 intracellular region and 2.6×104 fusion protein were obtained. Conclusion: The fusion protein containing polycystin-1 intracellular region is obtained and is helpful for preparing anti-polycystin-1 monoclonal antibody.
RESUMO
Objective: To clone hGLP-1 cDNA in the pBS SK(+/-)vector and construct the expression vector of pGEX-4T-3/hGLP-1cDNA to express GST-hGLP-1 fusion protein. Methods: The hGLP-1 cDNA was constructed by 6 synthetic oligonucleotides fragments, followed by the procedure of annealing and ligation with oligonucleotides fragments. The hGLP-1 cDNA was cloned into the pBS SK(+/-) vector, and was selected by α-complementation. It was confirmed by DNA sequening, then inserted into the MCS of the fusion expression vector pGEX-4T-3. The recombinant vector was transformed into E. coli TG1. Results: The recombinant plasmid DNA was digested with restrictive endonuclease BamHⅠand XhoⅠ. The result demonstrated that the hGLP-1 cDNA was successfully inserted into the pGEX-4T-3 vector and fusion protein GST-hGLP-1 had been expressed in SDS-PAGE. Conclusion： Expression of GST-hGLP-1 fusion protein can provide foundation for obtaining a larger quantity of recombinant hGLP-1 for experimental and clinic studies.
RESUMO
Objective: To obtain polycystin-1 intracellular region. Methods: cDNA of polycystin-1 intracellular region was generated by PCR and then cloned into pProEX Hta, which was prokaryotic expression vector. After verified by sequencing, the recombinant was transformed into E.coli host to express and purify the fusion protein by affinity chromatography. Results: 660 bp cDNA of polycystin-1 intracellular region and 2.6×104 fusion protein were obtained. Conclusion: The fusion protein containing polycystin-1 intracellular region is obtained and is helpful for preparing anti-polycystin-1 monoclonal antibody.
RESUMO
Objective: To clone hGLP-1 cDNA in the pBS SK(+/-)vector and construct the expression vector of pGEX-4T-3/hGLP-1cDNA to express GST-hGLP-1 fusion protein. Methods: The hGLP-1 cDNA was constructed by 6 synthetic oligonucleotides fragments, followed by the procedure of annealing and ligation with oligonucleotides fragments. The hGLP-1 cDNA was cloned into the pBS SK(+/-) vector, and was selected by α-complementation. It was confirmed by DNA sequening, then inserted into the MCS of the fusion expression vector pGEX-4T-3. The recombinant vector was transformed into E. coli TG1. Results: The recombinant plasmid DNA was digested with restrictive endonuclease BamHⅠand XhoⅠ. The result demonstrated that the hGLP-1 cDNA was successfully inserted into the pGEX-4T-3 vector and fusion protein GST-hGLP-1 had been expressed in SDS-PAGE. Conclusion： Expression of GST-hGLP-1 fusion protein can provide foundation for obtaining a larger quantity of recombinant hGLP-1 for experimental and clinic studies.