Use of the Caulobacter crescentus genome sequence to develop a method for systematic genetic mapping.

Journal of Bacteriology
Lisandra WestC Stephens

Abstract

The functional analysis of sequenced genomes will be facilitated by the development of tools for the rapid mapping of mutations. We have developed a systematic approach to genetic mapping in Caulobacter crescentus that is based on bacteriophage-mediated transduction of strategically placed antibiotic resistance markers. The genomic DNA sequence was used to identify sites distributed evenly around the chromosome at which plasmids could be nondisruptively integrated. DNA fragments from these sites were amplified by PCR and cloned into a kanamycin-resistant (Kan(r)) suicide vector. Delivery of these plasmids into C. crescentus resulted in integration via homologous recombination. A set of 41 strains containing Kan(r) markers at 100-kb intervals was thereby generated. These strains serve as donors for generalized transduction using bacteriophage phiCr30, which can transduce at least 120 kb of DNA. Transductants are selected with kanamycin and screened for loss of the mutant phenotype to assess linkage between the marker and the site of the mutation. The dependence of cotransduction frequency on sequence distance was evaluated using several markers and mutant strains. With these data as a standard, previously unmapped mutations were...Continue Reading

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Citations

Sep 11, 2007·Journal of Bacteriology·Sarah J ReisingerKathleen R Ryan
Oct 16, 2007·Journal of Bacteriology·Craig StephensUrs Jenal
Dec 19, 2006·Journal of Bacteriology·Craig StephensUrs Jenal
Dec 4, 2004·Journal of Bacteriology·Plamena Entcheva-Dimitrov, Alfred M Spormann
Feb 20, 2004·Journal of Bacteriology·Alison K HottesCraig Stephens
Apr 30, 2009·Molecular Microbiology·Andrea Möll, Martin Thanbichler
Apr 20, 2012·Molecular Microbiology·Patrick D CurtisYves V Brun
Jul 6, 2010·Molecular Microbiology·Cécile BerneYves V Brun
Aug 27, 2013·Microbiology·Carmen Fernandez-FernandezJustine Collier
Dec 20, 2005·Molecular Microbiology·Melanie L LawlerYves V Brun
Mar 15, 2006·Cell·Edgar HuitemaPatrick H Viollier
Dec 16, 2004·FEMS Microbiology Letters·Sunhee LeeWilliam R Jacobs
Nov 13, 2014·MBio·Heather A FeagaKenneth C Keiler
Mar 31, 2016·PLoS Genetics·Heather A FeagaKenneth C Keiler
Oct 28, 2017·Science·Courtney K EllisonYves V Brun
Aug 13, 2013·Journal of Bacteriology·Anastasiya A Yakhnina, Zemer Gitai
Feb 18, 2014·Journal of Bacteriology·Wolfgang StrobelMartin Thanbichler
Sep 4, 2019·Genetics and Molecular Biology·Alexy O ValenciaRodrigo S Galhardo
Mar 6, 2019·Journal of Bacteriology·Courtney K EllisonYves V Brun
May 8, 2019·Journal of Bacteriology·Nina I SulkowskiTanmay A M Bharat
Mar 10, 2017·BMC Evolutionary Biology·Roland Mathis, Martin Ackermann
Aug 24, 2017·Molecular Microbiology·Aleksandra ZielińskaMartin Thanbichler

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