植物乳植杆菌P-8连续传代过程中遗传稳定性分析

Genetic stability of Lactiplantibacillus plantarum P-8 during continuous subculturing

【背景】植物乳植杆菌(Lactiplantibacillus plantarum)P-8是一株具有优良益生特性的乳酸菌,探究其短期连续传代过程中的遗传稳定性对评估其加工生产的稳定性有着重要的指导作用。【目的】研究植物乳植杆菌P-8在37℃恒温环境下、MRS培养基中连续传代100代过程中的遗传稳定性。【方法】在MRS培养基、37℃恒温环境下将植物乳植杆菌P-8连续传代100代,测定不同代菌株(第0、25、50、75和100代)的菌体形态和碳水化合物利用能力,并利用二代、三代测序相结合的技术完成不同代菌株全基因组测序,通过比较基因组学综合分析其在连续传代100代过程中的遗传稳定性。【结果】植物乳植杆菌P-8在连续培养100代过程中,其菌体形态和碳水化合物利用能力均无明显变化。以植物乳植杆菌P-8的原始菌株基因组作为参考,比较分析了不同代菌株的基因组稳定性,发现菌株均有单核苷酸多态性(single nucleotide polymorphism,SNP)位点存在,但数量较少(SNP位点<21个)。不同代菌株基因组共线性良好,具有极高的相似性,且不同代菌株碳水化合物活性酶注释结果无显著差异(P>0.05)。【结论】植物乳植杆菌P-8在MRS培养基中、37℃恒温环境下连续传代100代期间具有良好的遗传稳定性,为该菌株的产业化应用奠定了遗传学基础。

[Background]Lactiplantibacillus plantarum P-8 is a lactic acid bacterium with excellent prebiotic properties.Exploring its genetic stability during short-term continuous subculturing helps to evaluate the stability of the cells in production.[Objective]To study the genetic stability of L.plantarum P-8 subcultured in MRS medium at 37℃for 100 generations.[Methods]We observed the cell morphology and measured the carbohydrate utilization ability of L.plantarum P-8 subcultured for 0,25,50,75,and 100 generations in MRS medium at 37℃.The whole genomes of cells subcultured for different generations were sequenced by the second-and third-generation sequencing.Comparative genomics was employed to comprehensively analyze the genetic stability of the cells during the 100 generations.[Results]The cell morphology and carbohydrate utilization ability of L.plantarum P-8 showed no significant changes during continuous subculturing for 100 generations.With the genome of the original cells as a reference,the genomic stability of different generations was compared.The cells of all the generations had single nucleotide polymorphism(SNP)sites,while the number of SNP sites was<21.The genomes of the cells at different generations had good collinearity and high similarity.The results of carbohydrate activity enzyme annotation showed no significant differences among different generations(P>0.05).[Conclusion]L.plantarum P-8 has good genetic stability during continuous subculturing for 100 generations in MRS medium at 37℃.This study laid a genetic foundation for the industrial application of this strain.