Single-copy Insertion of Transgenes in Caenorhabditis Elegans

Nat Genet. 2008 Nov;40(11):1375-83. doi: 10.1038/ng.248. Epub 2008 Oct 26.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Genetically Modified
  • Caenorhabditis elegans / cytology
  • Caenorhabditis elegans / genetics*
  • DNA Transposable Elements / genetics
  • Female
  • Gene Dosage*
  • Germ Cells
  • Green Fluorescent Proteins / metabolism
  • Injections
  • Male
  • Mutagenesis, Insertional / methods*
  • Transgenes / genetics*

Substances

  • DNA Transposable Elements
  • Green Fluorescent Proteins