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Chapter 31 : An Ordered Collection of Bacillus subtilis DNA Segments in Yeast Artificial Chromosomes
A collection of Bacillus subtilis chromosomal DNA fragments inserted in yeast artificial chromosomes (YACs) is represented in this chapter. YACs were obtained by random cloning of B. subtilis DNA, partially digested with EcoRI, in the pYAC4 vector. To order the YACs, we first used 66 previously cloned B. subtilis genes as hybridization probes. Next, the ends of the inserts carrying these genes were isolated. The ends were used as probes to identify clones with overlapping inserts. Iteration of the procedure resulted in the ordered collection. The identity of several segments was ascertained by transformation of B. subtilis mutants. Despite the relatively high redundancy of the random collection, four gaps remain in the ordered collection. Maximum gap size was estimated by hybridizing B. subtilis DNA cleaved with several restriction enzymes with the ends of segments bordering the gaps. The ordered collection should be useful for genetic studies in B. subtilis.