| Nisin is a small, 34-amino acid peptide produced by several strains of Lactococcuslactis,has a strong inhibition against many Gram-positive bacteria, includingfood-spoilage and harmful to human health pathogens such as Staphylococcus、Streptococcus and Listeria. As the most widely used high-efficient, innoxious naturalfood preservative, nisin has been widely applied in the preservation of meat, dairyproducts, eggs, fruits, beer brewing and it recently demonstrated broad applicationprospects in biomedical fields.In order to raise the level of nisin production, this paperfocused on the breeding of nisin producing strains, Lactococcus lactis.mutant strain MELN4 was obtained from Lactococcus lactis ATCC 11454 bytreatment with physical and chemical factors,such as ultraviolet, DES and LiC1. The nisinactivity of MELN4 was 2400 IU/mL, improved by 150% compared to Lactococcuslactis ATCC 11454.But after 6 generations,the nisin activity of MELN4 was only 1616IU/mL,which showed that the strain has not genetic stability.The Construction of high-yield nisin producing strain was investigated. The gene gadBencoding ghitamate decarboxylase from Ecoli TG1 was doned into Lactococcus lactis MELN4.ButEnzyme activity tests showed that the gadB gene in MELN4 was not active.The genes gadB and gadC encoding glutamate decarboxylase andglutamate-gamma-aminobutyrate antiporter were amplified from the chromosomal DNAof lactococcus lactis il1403, and then inserted into plasmid pMG36e to get a hybridplasmid pMG36egad. The hybrid plasmid pMG36egad were introduced into LactococcusIactis subsp, lactis (ATCC11454) by electroporation to get the recombinant bacteriaMELgad104.Enzyme activity tests showed that the gadCB gene in MEL4gad104 wasactively expressed.And the effect of expression gadCB gene on the fermentationperformance of MEL4gad104 was explored, results showed that the biomass and nisinactivity of the recombinant bacteria were increased by 24.1% and 19.7% respectively. |
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