Toward functional genomics in bacteria: Analysis of gene expression in Escherichia coli from a bacterial artificial chromosome library of Bacillus cereus

摘要

As the study of microbes moves into the era of functional genomics, there is an increasing need for mo- lecular tools for analysis of a wide diversity of microorgan- isms. Currently, biological study of many prokaryotes of agricultural, medical, and fundamental scientific interest is limited by the lack of adequate genetic tools. We report the application of the bacterial artificial chromosome (BAC) vector to prokaryotic biology as a powerful approach to address this need. We constructed a BAC library in Esche- richia coli from genomic DNA of the Gram-positive bacterium Bacillus cereus. This library provides 5.75-fold coverage of the B. cereus genome, with an average insert size of 98 kb. To determine the extent of hetcrologous expression of B. cereus genes in the library, we screened it for expression of several B. cereus activities in the E. coli host. Clones expressing 6 of 10 activities tested were identified in the library, namely, ampi- cillin resistance, zwittermicin A resistance, esculin hydrolysis, hemolysis, orange pigment production, and lecithinase activ- ity. We analyzed selected BAC clones genetically to identify rapidly specific B. cereus loci. These results suggest that BAC libraries will provide a powerful approach for studying gene expression from diverse prokaryotes.