Origin of the EcoRI Endonuclease G Fragment in the SP02 Bacteriophage Mutant Clh2 and Determination of the Temporal Order and Genomic Location of SP02 Bacteriophage Transcripts

Abstract

In developing Bacillus subtilis as a cloning host, phage for the host should be isolated and characterized. A promising phage that can be utilized as a cloning vector is bacteriophage SP02. SP02 has deletion mutants and the host-range for the phage has been extended to include Bacillus amyloliquefaciens and Bacillus natto. SP02 has been shown to have several unique endonuclease sites and, more importantly, there is a unique BglII endonuclease site in a genetically silent region of the genome. Deletion mutants for bacteriophage SP02 have been isolated. A host-range mutant of SP02, clh2 has an EcoRI digestion fragment that is foreign to the SP02 DNA. The first goal of this study was to determine the host with which SP02 DNA recombined to form the clh2 mutant. Initial findings from this study show some homology of biotinylated clh2 DNA to Bacillus amyloliquefaciens HSR chromosomal DNA. A second goal of this study was to determine the temporal order and genomic locations of SP02 transcripts. By hibridizing RNA transcripts, isolated temporally, to digested SP02 DNA, results from this study confirm a silent region in the SP02 genome, show temporal ordering of the SP02 transcripts, and show the genomic location of the SP02 transcripts.

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