Physical Mapping of the Synechococcus Sp. Strain PCC 7942 Chromosome
Ransom, Wendy D.
MetadataShow full item record
A partial physical map of the Synechococcus sp. strain PCC 7942 chromosome was constructed by the ordering of restriction fragments resolved with pulse field gel electrophoresis, cosmid-based walking using a preexisting cosmid library, and gene specific probings of pulse field gels. Cosmid-based walking on the right side of the rbcL locus was extended 60 kb by probing the library for related cosmids and by examining the restriction patterns of related cosmids. Problems of pseudo-linking cosmids to the existing map of the rbcL locus, and identification of cosmids in two different areas of the chromosome ended mapping at that locus. Chromosomal DNA was cleaved with Sfi I restriction endonuclease into 11 separable fragments by pulsed field gel electrophoresis. Five of the 11 fragments were linked together through cross hybridization with a gene-specific probe containing psbAIII, and linking cosmids from the library were identified on the fragments. The cosmids identified from hybridization of the fragments with the library revealed groups of cosmids shared between the fragments. These data indicated that the five fragments resulted from partial digestion with the enzyme, such that some of the fragments were overlapping subfragments of the others. A Synechococcus wox a gene-specific probe was hybridized with the cosmid library identifying cosmids containing the gene in the chromosome. One of the cosmids identified was mapped using the restriction endonucleases MluI, HindIII, BamHI, XbaI, and NotI to determine the position of the gene on the cosmid. The cosmids containing woxA were recognized by hybridization of the Sfi 9 fragment with the library and woxA was positioned within the Sfi 9 fragment through examining restriction patterns generated from Hind III / EcoRI double digest.