假单胞菌单基因甲基化位点的检测与自发表型变异分析

Single-gene DNA Methylation Sites Detection and Spontaneous Phenotypic Variation Analysis in Pseudomonas

重亚硫酸盐测序(BSP)技术是一种广泛应用的DNA甲基化检测技术,其检测分辨率达到单碱基水平,可用于短片段的高精度测序。利用BSP技术对DNA水平上不存在差异的绿针假单胞菌HT66及其荧光表型突变株HT66-FLUO的同一基因片段gacS并行测定,发现HT66-FLUO的gacS中存在2个甲基化位点,甲基化率分别为50%和100%,确定全局性调控基因gacS发生了甲基化修饰,从而影响了gacS的功能,导致HT66-FLUO自发表型变异。研究表明,BSP检测方法可应用于细菌单基因的甲基化位点分析,为细菌甲基化研究提供借鉴。

Bisulfite sequencing PCR(BSP)technology is a widely-used in DNA methylation detection.Its detection resolution can reach the single-base level and is suitable for high-precision sequencing of short fragments.In this study,gacS of Pseudomonas chlororaphis HT66 and its phenotype variant strain HT66-FLUO,which does not differ at the DNA level,were measured in parallel by BSP technology.It was found that there are two methylation-variable positions(MVPS)in the gacS of HT66-FLUO,with a methylation rate of 50% and 100%,respectively.It was confirmed that the global regulatory gene gacS is methylated,which affects the function of gacS and leads to HT66-FLUO spontaneous phenotypic variants.This study shows that the BSP detection method can be applied to the analysis on the methylation sites of single genes,providing a reference for the study of bacterial methylation.