Prokaryotic expression of antibiotic peptide Cecropin A gene and identification of expression products
10.3969/j.issn.1673-8225.2011.24.044
- VernacularTitle:抗菌肽Cecropin A基因原核表达及表达产物的鉴定
- Author:
Chen LEI
- Publication Type:Journal Article
- From:
Chinese Journal of Tissue Engineering Research
2011;15(24):4549-4552
- CountryChina
- Language:Chinese
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Abstract:
BACKGROUND: Cecropins are a kind of micromolecule protein with antibacterial activity. Eukaryotic cell-expressed or artificially synthesized Cecropins is characterized by low efficiency and high cost. OBJECTIVE: To clone and express an antibacterial peptide gene of Musca domestica Cecropin A, and to identify recombinant expression product. METHODS: Mature Musca domestica Cecropin A encoding nucleotide sequence was searched from the GenBank and amplified by RT-PCR. The gene of Musca domestica Cecropin A was cloned into prokaryotic expression vector pET32a and fused with gene of Thioredoxin (Trx) and expressed in E.coli BL2l (DE3). After induction by isopropyl-β-D-thiogalactoside, the sera of the immunized rabbits were collected after rabbits were immunized with the hemolymph of housefly larvae. Recombinant protein was identified by western blot analysis and N-[Tris(hydroxymethyl)methyl]glycine-sodium dodecylsulfate-polyacrylamide gel electrophoresis.RESULTS AND CONCLUSION: After induction by isopropyl-β-D-thiogalactoside, E.coli BL21 expressed mature Cecropin. Rabbit anti- housefly larvae sera, N-[Tris(hydroxymethyl)methyl]glycine-sodium dodecylsulfate-polyacrylamide gel electrophoresis and western blot analysis results confirmed that expression products were mature Cecropin. These suggest that prokaryotic expression system can be utilized to obtain natural mature Cecropin.
