| DNA cytosine methylation is a central epigenetic modification that has essential roles in the control of many critically important biological processes, including cell proliferation, differentiation, development, genomic imprinting and regulation of gene expression. With progresses in study on DNA methylation, detection techniques of DNA methylation have been developed to meet requirements of various researches.This article has established an improved bisulfite sequencing technique for analysis of MutL-homologue 1（MLH1） promoter methylation level of mismatch repair gene in Arabidopsis thaliana. After the genomic DNA in Arabidopsis thaliana was digested, purified, and then modified with bisulfite liquid, the PCR products were cloned into the pEASY-T1 vector. 15 positive clones were randomly selected from each sample and then sequenced. The results showed that compared with the traditional method of methylation analysis,the improved method modified the digested DNA completely, reduced the modification time greatly, and improved the specificity, stability and repeatability of the PCR products. MLH1 promoter methylation level was 27.3% instead of 45.6%. The improved bisulfite sequencing technique avoided misidentification at non-methylated sites, and would provide a better means of research for analysis of DNA methylation in plant. |
