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PMID: 9321655 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Target choice determinants of the Tc1 transposon of Caenorhabditis elegans.

Nucleic acids research ·Vol. 25 ·No. 20 ·1997-10-15 ·Pages 4041-7

Ketting RF, Fischer SE, Plasterk RH

Abstract

The Tc1 transposon of Caenorhabditis elegans always integrates into the sequence TA, but some TA sites are preferred to others. We investigated a TA target site from the gpa-2 gene of C.elegans that was previously found to be preferred (hot) for Tc1 integration in vivo . This site with its immediate flanks was cloned into a plasmid, and remained hot in vitro , showing that sequences immediately adjacent to the TA dinucleotide determine this target choice. Further deletion mapping and mutagenesis showed that a 4 bp sequence on one side of the TA is sufficient to make a site hot; this sequence nicely fits the previously identified Tc1 consensus sequence for integration. In addition, we found a second type of hot site: this site is only preferred for integration when the target DNA is supercoiled, not when it is relaxed. Excision frequencies were relatively independent of the flanking sequences. The distribution of Tc1 insertions into a plasmid was similar when we used nuclear extracts or purified Tc1 transposase in vitro , showing that the Tc1 transposase is the protein responsible for the target choice.

MeSH Terms
Animals Base Composition Base Sequence Binding Sites Caenorhabditis elegans/chemistry,genetics DNA/chemistry,metabolism DNA Transposable Elements DNA, Superhelical/metabolism DNA-Binding Proteins/metabolism Gene Deletion Mutagenesis, Site-Directed Nucleotidyltransferases/metabolism Plasmids Polymerase Chain Reaction Structure-Activity Relationship Transposases
Chemicals
DNA Transposable Elements DNA, Superhelical DNA-Binding Proteins DNA Nucleotidyltransferases Tc1 transposase Transposases
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Ketting R F
Division of Molecular Biology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands.
Fischer S E
Plasterk R H
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/25.20.4041
Published
1997-10-15
Pages
4041-7
Language
English
Region
England
NLM ID
0411011
PMCID
PMC147011
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/25.20.4041 PMC Full Text View journal details
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