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| Construction of prokaryotic expression vector and preparation of mouse bile salt dependent lipase protein |
| YANG Yi SONG Huanlei QIAN Linxi |
| Xinhua Hospital Affiliated to School of Medicine of Shanghai Jiaotong University Shanghai Institute for Pediatric Research Shanghai Key Laboratory of Pediatric Gastroenterology and Nutrition, Shanghai 200092, China |
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Abstract Objective To construct a prokaryotic expression system for mBSDL in E. coli BL21 (DE3) cells and prepare the active mBSDL protein. Methods mBSDL cDNA was amplifed via PCR and inserted into the prokaryotic expression vector pET-28b. The expression of recombinant protein was induced by IPTG in E. coli BL21 cells. Then it was purified via His Trap FF crude purification system and refolded via dialysis system. The specificity of mBSDL protein was analyzed by Western blot. Results The prokaryotic expression vector pET-28b-mBSDL expressing mBSDL protein was successfully constructed and transformed into E. coli BL21(DE3) cells. The mBSDL protein was present as insoluble inclusion bodies in the bacterial extract. Western blot reveled that the purification of this recombinant protein was successful. Conclusion mBSDL protein in E. coli prokaryotic expression system is established successfully. The study provides some foundation for the further analysis of mBSDL protein.
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