Transposon-mediated restriction mapping of the Bacillus subtilis chromosome.
Analysis of chromosomal DNA depends upon a knowledge of the locations of restriction sites over several thousand kilobases (kb). However determination of even a subset of these sites can be time-consuming, and it can be difficult to link genetic and physical maps. We describe here a significant improvement which can be used in concert with genetically mapped chromosomal insertions. The circular chromosome of Bacillus subtilis 168 was physically examined on contour-clamped homogeneous electric field (CHEF) gels using the restriction enzyme NotI. Restriction mapping of the 4.7-megabase (Mb) DNA was accomplished using a novel technique involving the transposon Tn917, which linked the genetic and physical maps and also significantly increased the rate at which this was performed. The DNA of 54 strains which contained Tn917 at genetically determined locations was cleaved with NotI and used to determine the approximate positions of 31 restriction fragments with sizes between 45 kb and 290 kb, totalling 3589 kb. This information should greatly assist in the construction of a more detailed map using standard methodology.
Ventra
L
L
Department of Biochemistry, Unviersity of Sydney, NSW, Australia.
Weiss
A S
AS
D001412:Bacillus subtilis / Q000235:genetics*; D015139:Blotting, Southern; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D015342:DNA Probes; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D015183:Restriction Mapping
D015342:DNA Probes; D004251:DNA Transposable Elements; D004269:DNA, Bacterial