Rat HMGB1 protein in prokaryotic expression and purification.docVIP

 PAGE \* MERGEFORMAT 13 Rat HMGB1 protein in prokaryotic expression and purification Author: Yang Hui Liu Zhao-fu Feng Jiangnan Social Zhang Yun Yang catkins [Keywords:] high mobility group box protein 1; prokaryotic expression vector; purification Abstract Objective: To clone mouse HMGB1 gene, construct prokaryotic expression vector induced expression in E. coli and purified to obtain His-HMGB1 fusion protein. Methods: The LPS stimulation of RAW264.7 cells, extracted total RNA, was amplified by RT-PCR with the purpose of HMGB1 fragment was cloned into pMD-19T vector, then subcloned to contain pelB peptide and the His-tag peptide Efficient expression vector pET-26b (), transformed into E. coli BL21 (DE3), induced by IPTG Across SDSidentification of target protein expression, using nickel-chelating affinity chromatography agarose gel containing purified His-tagged peptide the target protein. Results: The PCR amplification of HMGB1 size of 648 bp gene fragment was successfully constructed, the target protein fusion expression vector, induced expression and purification, received about 30 kD fusion protein. Conclusion: The Construction of HMGB1 fusion expression vector to obtain purified fusion protein His-HMGB1, for further research laid the foundation for its biological function. Keywords: high mobility group box protein 1; prokaryotic expression vector; purification Mouse HMGB1 Prokaryotic Expression and Purification Abstract Objective: To clone mouse High mobility group box 1 (HMGB1) gene and construct the prokaryotic expression vector, to induce and purify the expressed fusion protein HMGB1.Methods: The total RNA was extracted from mouse RAW264.7 cells stimulated by LPS.The sequence including the whole length of HMGB1 was amplified by RT-PCR and inserted into pMD-19T.The combinant vector was used as a template for PCR which was cloned into vector pMD-19T, then subcloned into expression vector pET-26b () with pelB signal sequence and His-Taq sequence.Aft