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Updated: Jun 14, 2026
Protocols for Implementing an Escherichia coli Based TX-TL Cell-Free Expression System for Synthetic Biology
Published on: September 16, 2013
[Expression of human TFF3 in Escherichia coli]
Rui Lu1, Xin Wang, Jian-Ping Chen
1Department of Parasitology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu, China.
Objective:
To construct recombinant human TFF3 prokaryotic expressing plasmid, express it in E. coli and identify the expressed protein.
Methods:
The cDNA for mature peptide of TFF3 was amplified by RT-PCR from RNA of human colon tissue and inserted into the MCS of the prokaryotic expressing plasmid pET32a (+). Then TFF3 was expressed as a fusion protein by IPTG induction. The recombinant protein was determined by SDS-PAGE and Western blot with a rabbit anti-TFF3 polyclonal antibody.
Results:
Sequencing result indicated that the obtained TFF3 fragment was inserted into plasmid pET32a (+) successfully and the sequence was correct. The expression level of the fusion protein was highest after 6h induction with 1 mmol/L IPTG. The result of Western blot demonstrated that the relative molecular mass of recombinant protein was about 24 x 10(3) and the protein had good antigenicity and specificity.
Conclusion:
The expression plasmid pET32a-TFF3 was constructed and expressed successfully. This study will provide a substantial basis for further study of human TFF3.
