同源重组法构建副干酪乳杆菌组氨酸蛋白激酶基因缺失突变株

发布时间：2018-05-14 00:41

本文选题：副干酪乳杆菌 + 细菌素　；参考：《食品科学》2017年12期

【摘要】：构建副干酪乳杆菌(Lactobacillus paracasei)组氨酸蛋白激酶prcK基因缺失突变株,为prcK基因功能研究提供实验工具。采用同源重组技术构建质粒p KLKRT(含prcK::Tet~r基因),将其电转化进入L.paracaseiHD1.7中,使prcK::Tet~r基因同源重组到L.paracasei HD1.7的染色体上,通过四环素耐药、氨苄青霉素敏感等特性筛选出含有prcK::Tet~r基因的新L.paracasei HD1.7。结果表明,经聚合酶链式反应(polymerase chain reaction,PCR)验证及酶切验证后确定质粒p KLKRT构建成功,并成功转化入L.paracasei HD1.7中,经PCR确认L.paracasei HD1.7 prcK基因缺失突变株构建成功。该突变株产生的细菌素效价比出发菌株低23.61%。采用同源重组方法成功构建L.paracasei HD1.7 prcK基因缺失突变株,为研究L.paracasei HD1.7群体效应相关基因的分子机理奠定基础。
[Abstract]:The mutant of histidine protein kinase (prcK) of Lactobacillus paracia paracae was constructed to provide experimental tools for the study of prcK gene function. The plasmid pKLKRT( containing prcK::Tet~r gene) was constructed by homologous recombination technique, which was electrotransformed into L.paracaseiHD1.7, and the prcK::Tet~r gene was homologous recombined into the chromosome of L.paracasei HD1.7. The new L.paracasei HD1.7 containing prcK::Tet~r gene was screened by tetracycline resistance and ampicillin sensitivity. The results showed that the plasmid p KLKRT was successfully constructed and transformed into L.paracasei HD1.7 by polymerase chain reactionation (PCR) and restriction endonuclease assay (PCR), and the L.paracasei HD1.7 prcK gene deletion mutant was successfully constructed by PCR. The bacteriocin titer of the mutant was 23.61 lower than that of the original strain. The homologous recombination method was used to construct L.paracasei HD1.7 prcK gene deletion mutants, which laid a foundation for studying the molecular mechanism of L.paracasei HD1.7 population effect-related genes.
【作者单位】：黑龙江大学生命科学学院微生物省高校重点实验室;黑龙江大学农业微生物技术教育部工程研究中心;
【基金】：国家自然科学基金面上项目(31470537;31570492) 黑龙江省高等学校科技创新团队(农业微生物发酵技术)项目(2012td009)
【分类号】：TS201.3

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