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PMID: 1735708 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Chromosome transfer in Rhodobacter sphaeroides: Hfr formation and genetic evidence for two unique circular chromosomes.

Journal of bacteriology ·Vol. 174 ·No. 4 ·1992-02-00 ·Pages 1135-45

Suwanto A, Kaplan S

Abstract

A 600-bp oriT-containing DNA fragment from the Rhodobacter sphaeroides 2.4.1 S factor (oriTs) (A. Suwanto and S. Kaplan, J. Bacteriol. 174:1124-1134, 1992) was shown to promote polarized chromosomal transfer when provided in cis. A Kmr-oriTs-sacR-sacB (KTS) DNA cassette was constructed by inserting oriTs-sacR-sacB into a pUTmini-Tn5 Km1 derivative. With this delivery system, KTS appeared to be randomly inserted into the genome of R. sphaeroides, generating mutant strains which also gained the ability to act as Hfr donors. An AseI site in the Kmr cartridge (from Tn903) and DraI and SnaBI sites in sacR-sacB (the levansucrase gene from Bacillus subtilis) were employed to localize the KTS insertion definitively by pulsed-field gel electrophoresis. The orientation of oriTs at the site of insertion was determined by Southern hybridization analysis. Interrupted mating experiments performed with some of the Hfr strains exhibited a gradient of marker transfer and further provided genetic evidence for the circularity and presence of two chromosomal linkage groups in this bacterium. The genetic and environmental conditions for optimized mating between R. sphaeroides strains were also defined. The results presented here and our physical map of the R. sphaeroides 2.4.1 genome are discussed in light of the presence of two chromosomes.

MeSH Terms
Chromosomes, Bacterial/metabolism Conjugation, Genetic/genetics DNA Replication/genetics DNA, Circular/genetics Electrophoresis, Gel, Pulsed-Field Genetic Vectors/genetics Rhodobacter sphaeroides/genetics
Chemicals
DNA, Circular
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Suwanto A
Department of Microbiology and Molecular Genetics, University of Texas Health Science Center, Houston 77225.
Kaplan S
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.174.4.1135-1145.1992
Published
1992-02-00
Pages
1135-45
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC206406
Subset
IM
Grants
NIGMS NIH HHS · GM15590 · United States
NIGMS NIH HHS · GM31667 · United States
External Links
PubMed source DOI: 10.1128/jb.174.4.1135-1145.1992 PMC Full Text View journal details
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