Expression, Purification and Characterization of Populus tomentosa PtoeIF5A4 in Prokaryote

Abstract

Abstract: To obtain the PtoeIF5A4 protein of poplar with high purity in a large scale, its full length CDS was amplified by PCR using pGEM-T Easy-PtoeIF5A4 as template and verified by sequencing, and then was inserted into pET28a vector containing histidine. The recombinant pET28-PtoeIF5A4 plasmid was obtained and transformed into E. coli BL21(DE3). The recombinant fusion protein was produced by culturing the transformed E. coli BL21(DE3)with subsequent IPTG induction, and was observed predominantly in the supernatant of cellular extracts when cells were cultured at 28℃ and induced with 0.1 mmol/L IPTG for 4 h. The fusion protein was purified from the soluble fraction using a column of Ni²⁺ Chelating Sepharose Fast Flow, and proofed molecular mass was 18 kD. The anti-His monoclonal antibody recognized the protein, which indicated that it was PtoeIF5A4 recombinant protein.

Key words: Populus tomentosa, PtoeIF5A4, Prokaryotic expression, Protein, Purification

Zhang Jiewei, Guan Yang, Zhu Dan, Chen Yajuan, Ding Liping, Wang Hongzhi, Wei Jianhua. Expression, Purification and Characterization of Populus tomentosa PtoeIF5A4 in Prokaryote[J]. Biotechnology Bulletin, 2014, 0(5): 83-87.

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