Comprehensive identification of conditionally essential genes in mycobacteria
Comprehensive identification of conditionally essential genes in mycobacteria
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DOI：
10.1073/pnas.231275498
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发表时间：
2001-10-23
影响因子：
11.1
通讯作者：
Rubin, EJ
中科院分区：
文献类型：
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作者：
Sassetti, CM;Boyd, DH;Rubin, EJ
越来越多的微生物基因组已经被完全测序，并且所鉴定的基因基于其与已知功能的基因的同源性而被分类。然而，大量基因的功能不能仅在此基础上确定。在这里，我们描述了一种技术，转座子位点杂交（TraSH），它允许通过识别在不同条件下生长所需的完整基因集的快速功能表征。TraSH将高密度插入诱变与突变体库的微阵列作图相结合。我们通过使用基于水手的转座子和有效的噬菌体转导，在分枝杆菌中建立了大量独立的转座子突变体库。通过使用TraSH，我们已经定义了牛分枝杆菌卡介苗在最小但不丰富的培养基上生长所需的一组基因。鉴定了已知和未知功能的基因。在已知功能的基因中，几乎所有的基因都参与氨基酸的生物合成。TraSH是一种对基因功能进行分类的有力方法，应该适用于各种微生物。
An increasing number of microbial genomes have been completely sequenced, and the identified genes are categorized based on their homology to genes of known function. However, the function of a large number of genes cannot be determined on this basis alone. Here, we describe a technique, transposon site hybridization (TraSH), which allows rapid functional characterization by identifying the complete set of genes required for growth under different conditions. TraSH combines high-density insertional mutagenesis with microarray mapping of pools of mutants. We have made large pools of independent transposon mutants in mycobacteria by using a mariner-based transposon and efficient phage transduction. By using TraSH, we have defined the set of genes required for growth of Mycobacterium bovis bacillus Calmette-Guerin on minimal but not rich medium. Genes of both known and unknown functions were identified. Of the genes with known functions, nearly all were involved in amino acid biosynthesis. TraSH is a powerful method for categorizing gene function that should be applicable to a variety of microorganisms.