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Single-copy insertion of transgenes in Caenorhabditis elegans.

ABSTRACT: At present, transgenes in Caenorhabditis elegans are generated by injecting DNA into the germline. The DNA assembles into a semistable extrachromosomal array composed of many copies of injected DNA. These transgenes are typically overexpressed in somatic cells and silenced in the germline. We have developed a method that inserts a single copy of a transgene into a defined site. Mobilization of a Mos1 transposon generates a double-strand break in noncoding DNA. The break is repaired by copying DNA from an extrachromosomal template into the chromosomal site. Homozygous single-copy insertions can be obtained in less than 2 weeks by injecting approximately 20 worms. We have successfully inserted transgenes as long as 9 kb and verified that single copies are inserted at the targeted site. Single-copy transgenes are expressed at endogenous levels and can be expressed in the female and male germlines.

SUBMITTER: Frokjaer-Jensen C

PROVIDER: S-EPMC2749959 | biostudies-literature | 2008 Nov

REPOSITORIES: biostudies-literature

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Single-copy insertion of transgenes in Caenorhabditis elegans.

Frøkjaer-Jensen Christian C Davis M Wayne MW Hopkins Christopher E CE Newman Blake J BJ Thummel Jason M JM Olesen Søren-Peter SP Grunnet Morten M Jorgensen Erik M EM

Nature genetics 20081026 11

At present, transgenes in Caenorhabditis elegans are generated by injecting DNA into the germline. The DNA assembles into a semistable extrachromosomal array composed of many copies of injected DNA. These transgenes are typically overexpressed in somatic cells and silenced in the germline. We have developed a method that inserts a single copy of a transgene into a defined site. Mobilization of a Mos1 transposon generates a double-strand break in noncoding DNA. The break is repaired by copying DN ...[more]

PMID: 18953339

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