Genome Reduction And Phenotypic Identification Of Lactococcus Lactis

With a crucial role in dairy and health industries,Lactococcus lactis is a GRAS(Generally Regarded as Safe)microorganism.Features of L.lactis such as lack of lipopolysaccharides and have few secreted proteins could make it to be an excellent expression host.However,an efficient genetic engineering system for L.lactis is still missing.In this study,a convenient gene deletion system was established and genome reduction mutants were constructed based on L.lactis NZ9000-3.Besides,the phenotypes and causes of mutants were analyzed in-depth.More detailed work is summarized as followed.1.Convenient and efficient system for large-scale genome deletion in L.lactis through the application of visually selectable marker was provided,which could be helpful for rapid genome streamlining and generation of restructured L.lactis strains that can be used as cell factories.2.Accounting for 8.566% in L.lactis NZ9000 genome was deleted sequentially.Compared with the parent strain,the mutant L.lactis NZ9000-5 showed better growth characteristics,transformation,carbon metabolic capacity,and amino acids biosynthesis.The results of this study indicated that further genome refinements and reductions in L.lactis could eventually generate a significantly simplified strain that could contribute to broadening the use of the bacterium.3.The fragment transferred mutant L.lactis NZ9000-6T obtained in the process of L.lactis NZ9000 genome reduction has an excellent growth phenotype,and the cause of the growth phenotype was also explored in this study.In summary,a high-throughput screening gene editing system was established for the construction of L.lactis chassis cells in this study.The genome reduction strains of L.lactis NZ9000,obtained with excellent phenotype,have broad prospects for microbial industrial application.