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PMID: 2165471 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Characterization of insertion sequence IS946, an Iso-ISS1 element, isolated from the conjugative lactococcal plasmid pTR2030.

Journal of bacteriology ·Vol. 172 ·No. 8 ·1990-08-00 ·Pages 4151-60

Romero DA, Klaenhammer TR

Abstract

The self-transmissible plasmid pTR2030 mobilized nonconjugative heterologous cloning vectors pGK12 (Cmr Emr) and pSA3 (Emr) at frequencies of 10(-5) to 10(-6) per input donor. Transconjugants harbored a 51- or 58-kilobase (kb) plasmid not found in the parental strains that cotransferred at high frequency with Cmr Emr and pTR2030-encoded phage resistance (Hsp+) in second-round matings (10(-1) per input donor). Restriction endonuclease mapping and DNA-DNA hybridization identified the 51- to 58-kb plasmids as pTR2030::vector cointegrates. Examination of four cointegrates indicated that pGK12 and pSA3 had inserted within two locations on pTR2030. Resolution of the cointegrates generated vector derivatives containing a 0.8-kb insert of pTR2030 DNA. Restriction analyses of several resolution plasmids indicated that the 0.8-kb element had inserted into various positions within pGK12 and pSA3 and in certain cases had inactivated the Cmr or Emr marker of pGK12. A conjugative mobilization assay demonstrated that the 0.8-kb element, designated IS946, mediated transpositional recombination. Nucleotide sequence determination identified IS946 as an 808-base-pair (bp) insertion sequence sharing ca. 96% homology with lactococcal insertion sequence ISS1. IS946 differed by 27 and 31 bp from ISS1S and ISS1T, respectively, and in 2 of 226 amino acids in the deduced sequence of the putative transposase. IS946 has perfect 18-bp terminal inverted repeats, identical to ISS1, and similarly generated 8-bp direct repeats of the target site upon insertion.

MeSH Terms
Amino Acid Sequence Base Sequence Cloning, Molecular Conjugation, Genetic DNA Transposable Elements Escherichia coli/genetics Lactococcus lactis/genetics Molecular Sequence Data Phenotype Plasmids R Factors Restriction Mapping Sequence Homology, Nucleic Acid
Chemicals
DNA Transposable Elements
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Romero D A
Department of Microbiology, Southeast Dairy Foods Research Center, North Carolina State University, Raleigh 27695.
Klaenhammer T R
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.172.8.4151-4160.1990
Published
1990-08-00
Pages
4151-60
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC213237
Subset
IM
External Links
PubMed source DOI: 10.1128/jb.172.8.4151-4160.1990 PMC Full Text View journal details
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