大肠杆菌质粒解离后致死基因分布及多态性分析

Distribution and polymorphisms of host killing genes in the plasmids metagenome of Escherichia coil

目的通过对来自小同年份大肠杆菌质粒DNA序列进行商通量测序，从质粒基因组水平分析大肠杆菌质粒DNA所携带的解离后致死基因的种类、数量以及多态性。方法收集4年临床分离非重复的320株大肠杆菌。菌株分为两个部分，碱裂解法提取全部质粒DNA，Solexa测序获得大规模的短序列。采用比较基因组学方法分析两个样本所含的解离后致死基因类型及丰度的差异，研究解离后致死基因存在的核苷酸多态位点。结果测序法获得两批数据，E1短序列总数为11077768，叮以定位到参照序列上为5019（0．045％）。E2短序列总数为9377792，可以定位到参照序列上是4952（0．053％）。两个样本中共有9个解离后致死基因型，其中hok基阕丰度最高。5个基因型共发现29个单核苷酸多态性（SNPs）位点，约33％为非同义突变。同一个位点的A、G二核苷酸多态现象常见。结论已知的大肠杆菌解离后致死基因的9个基因型，在温州地区分离的大肠杆菌质粒上均有发现，其中hok基因出现频率最高，共5个基因型存在多态性。

Objective To investigate the genotypes of host killing genes and their single nucleotide polymorphisms （SNPs）. Methods Three hundred and twenty strains of Escheriehia cull that collected from the First Affiliated Hospital of Wenzhou Medical College were analyzed. The first sample （ E1 ） contains 160 strains isolated during the years from 2002 to 2003. The second sample （E2） contains 160 strains coveting the years from 2008 to 2009. The plasmids of Escherichia cull were extracted by alkaline lysis method. Solexa/lllumina sequencing technology was used to sequence plasmids metagenome. Solexa Genome Analysis System and Soap programs were used to analyze gene distribution, SNPs and lineage-specific mutations. Resuits 11 077 768 reads were generated and 0. 045% of them can map to the reference sequences from E1 sample. Whereas 9 377 792 leads were generated and 0. 053% of which mapped to the reference from E2 sample. There are nine host killing genes identified in the two samples, of which hok gene is the most prevalent. A total of 29 SNP sites dispersed in five genes of the two samples. Approximately 33% of them were non-synonymous mutations. One position of A and G is the most prevalent polymorphism. Conclusion The known nine genotypes of host killing genes were all identified in plasmids of Escherichia cull in Wenzhou. hok gene showed the highest,hest frequencv. There were SNPs in five genotvnes.