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Tropical Journal of Pharmaceutical Research
Pharmacotherapy Group, Faculty of Pharmacy, University of Benin, Benin City, Nigeria
ISSN: 1596-5996
EISSN: 1596-5996
Vol. 16, No. 4, 2017, pp. 811-818
Bioline Code: pr17103
Full paper language: English
Document type: Research Article
Document available free of charge

Tropical Journal of Pharmaceutical Research, Vol. 16, No. 4, 2017, pp. 811-818

 en Simple and efficient expression of codon-optimized mouse leukemia inhibitory factor in Escherichia coli check for this species in other resources
Niu, Dewei; Wu, Qiuhong; Huang, Luyuan; Yang, Enze; Huang, Meiyan; Liu, Yong & Wang, Feng

Abstract

Purpose: To obtain a higher yield of mouse leukemia inhibitory factor to maintain the proliferation potential of pluripotent stem cells at a low cost.
Methods: A method was designed to produce recombinant mLIF protein (rmLIF) in Escherichia coli. Through analysis of rmLIF sequence, it was found that rare codons were interspersed. After mutation from rare codons to Escherichia coli (E. coli) preferred ones were selected, the mutated gene mLIFm was cloned into pET15b vector. The pET15b-mLIFm was then transformed into Rosetta-gami strain and induced with optimal conditions at 18 oC for 16 h. Mass spectrometry was carried out to identify the peptides.
Results: After purification, the yield of the codon-optimized rmLIFm was 141 mg/L, compared with 110 mg/L for the original rmLIF. Mass spectral analysis showed the presence of four major peptides each with an intensity > 10 % at m/z 1031.57, 1539.82, 1412.01 and 2229.10 in mLIFm, respectively. His-tagged rmLIFm fusion protein displayed the potential to maintain the morphology of undifferentiated mouse embryonic stem cells (mESCs), which were positive for mESCs markers (Oct-4, Nanog, Sox-2, stage-specific embryonic antigen-1).
Conclusion: The findings provide a means to produce mLIF in a short, useful, cost-effective and environmentally-friendly manner, and thus lays a foundation for further studies of mLIF.

Keywords
Leukemia inhibitory factor, Mutated gene, Protein expression, Purification, Stem cells, Peptides, Escherichia coli

 
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