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Mostly probiotic lactic acid bacteria (lactic acid bacteria), is at this stage studied a class of engineering strain. Lactic acid bacteria used in the food industry is generally considered to be safe, and therefore the research and development of lactic acid bacteria food-grade carrier - receptor system, discover important Cloning and expression of genes in lactic acid bacteria, lactic acid bacteria in molecular biology has become hot. From the separation of lactic acid bacteria in fermented foods, and then extract high purity plasmid, select the appropriate plasmid Sequencing and analysis of the lactic acid bacteria plasmid and host matching capabilities to study. Firstly, pickles and yogurt were isolated eight strains of lactic acid bacteria, Gram staining were positive, non-spore, catalase test was negative, facultative anaerobic growth of 5% CO2 conditions, metabolites lactic acid, bromocresol purple to turn yellow, no gas, and salt-tolerant growth. Identified by the 16S rDNA, six Lactobacillus plantarum, two Streptococcus thermophilus. Method for extracting natural lactic acid bacterium plasmid, and found and methods (O'Sullivan create Method), Method II (collection of lactic acid bacteria and the cleavage method established by O'Sullivan et plasmid was extracted and recovered in accordance with colorectal subtilis plasmid extraction method) compared to the use of tris (cell lysate treated with lysozyme, and then use Beijing TIANGEN plasmid extraction kit for extraction of plasmid) resulting plasmid strip more clear, bright better extraction . On the method for further optimization to determine the optimum concentration of lysozyme in the extraction process 20mg/mL, the optimum treatment time for 30min, and toxic substances while avoiding the use of ethidium bromide. From five lactic acid bacteria plasmid extraction, and select laboratory isolated from Kimchi Lactobacillus plantarum S1 extract about 2 kb the plasmid pLPS1 as the research object, the pLPS1 sequenced, bioinformatics the sequences were analyzed. The plasmid gene is 2117bp, G C content of 38.03%. From 267 to 1448bp open reading frame ORF1 encodes 393 amino acids. The theoretical molecular weight of the encoded protein is 44.3kDa. Encoding the protein secondary structure prediction, wherein 44.27% of the total amount of protein alpha-helical structure, extension of chain accounted for 7.38%, 4.58% beta-corner, irregular curl accounted for 43.77%. About a 20 amino acid protein sequence of the N-terminal hydrophobic region, the analysis shows that it has the function of the signal peptide. On the plasmid pMG36e electrical conversion conditions. Lactobacillus plant as host bacteria CGMCC1.555, the optimal conditions for transforming: Operating voltage 2.25kV, glycine concentration of 2.95%, cultured at 3.96h, under this condition, the power-conversion efficiency reaches the predicted maximum 1909cfu/μg DNA; Lactococcus lactis subsp CICC 6060 as the host bacteria, the optimum conditions for the electrical conversion: Operating voltage 2.38kV, glycine concentration of 2.68%, cultured at 6.19h, under this condition, the power-conversion efficiency prediction Max 1375cfu / μg DNA.
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