THE COMPLETE GENOME SEQUENCE AND ANALYSIS OF M SMEGMATIS

实验室菌株H37Rv和临床毒株CDC1551两株结核分枝杆菌DNA序列的完成和注释，将为分枝杆菌和结核病研究界研究结核分枝杆菌的传染性、毒力、定植、致病性以及药敏和耐药机制提供前所未有的资源。尽管基因组序列提供了丰富的信息，但这种生物体的生长速度极慢，毒性极强，因此处理结核分枝杆菌及其基因操作是一项艰巨的任务。为开发耻垢分枝杆菌（一种生长迅速的非病原体）作为结核分枝杆菌的遗传替代物，人们付出了巨大的努力。通过这些努力，分枝杆菌研究人员有可能以与大肠杆菌在更大的微生物群落中使用的方式大致相同的方式利用耻毛分枝杆菌。获得耻垢分枝杆菌的完整基因组序列将是分枝杆菌研究的一个里程碑。除了为其作为重组方法的遗传系统的继续发展提供宝贵的信息外，该生物还为研究分枝杆菌生物学的独特方面提供了有价值的模型，包括细胞壁生物合成，生长速率调节，药物敏感性，蛋白质分泌，基因调节等。基因组研究所开发了一种具有成本效益和效率的微生物基因组测序方法，该方法已用于对七个微生物基因组进行测序和组装。耻垢分枝杆菌mc2155的基因组将使用这种全基因组霰弹枪策略进行测序。将准备大小随机插入文库，并获得足够数量的随机霰弹枪序列，以达到8倍的覆盖率。基因组的组装将使用TIGR组装软件的2.0版本完成，并将通过研究所开发的各种基于PCR的技术获得封闭。基因组的注释将采用多种计算机技术来识别开放的阅读框，并通过对管理数据库的相似性搜索来识别已知功能的蛋白质。额外的分析将确定诸如输出蛋白、tRNA和rRNA基因、重复序列、启动子序列和核糖体结合位点等特征。这些数据将通过万维网上的TIGR微生物数据库（www.tigr.org）提供给研究界。

The completion and annotation of the DNA sequence from two strains of M. tuberculosis, the laboratory strain H37Rv and the virulent clinical isolate CDC1551 will provide the mycobacteria and tuberculosis research communities with an unprecedented resource for investigating the infectivity, virulence, colonization, pathogenicity, and the mechanisms of drug- sensitivity and resistance of M. tuberculosis. Despite the abundance of information provided by the genomic sequence, the exceedingly slow growth rate and highly virulent nature of the organism make working with M. tuberculosis and its genetic manipulation formidable tasks. A great deal of effort has gone into developing M. smegmatis, a fast-growing non-pathogen, as a genetic surrogate for M. tuberculosis. Through these efforts, it is possible for the mycobacterial researcher to utilize M. smegmatis in much the same manner as E. coli is used in the greater microbiological community. Obtaining the complete genome sequence of M. smegmatis will represent a milestone in mycobacterial research. In addition to providing invaluable information for continuing its development as a genetic system for recombinant methodologies, the organism provides a valuable model for studying unique aspects of mycobacterial biology, including cell wall biosynthesis, regulation of growth-rate, drug-sensitivity, protein secretion, gene regulation, etc. The Institute for Genomic Research has developed a cost effective and efficient approach to microbial genome sequencing that has been used to sequence and assemble seven microbial genomes. The genome of M. smegmatis strain mc2155 will be sequenced using this whole genome shotgun strategy. Small and large insert random libraries will be prepared and a sufficient number of random shotgun sequences to reach 8-fold coverage will be obtained. Assembly of the genome will be done using the version 2.0 of the TIGR Assembly software and closure will be obtained with a variety of PCR based techniques developed at The Institute. Annotation of the genome will employ a variety of computer techniques for identifying open reading frames and identifying proteins of known function through similarity searches against curated databases. Additional analyses will identify features such as exported proteins, tRNA and rRNA genes, repeated sequences, promoter sequences and ribosome binding sites. The data will be made available to the research community through the TIGR Microbial Database on the World Wide Web (www.tigr.org).