Single-copy insertion of transgenes in Caenorhabditis elegans

  • Frøkjær-Jensen C
  • Wayne Davis M
  • Hopkins C
 et al. 
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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.

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Authors

  • Christian Frøkjær-Jensen

  • M. Wayne Davis

  • Christopher E. Hopkins

  • Blake J. Newman

  • Jason M. Thummel

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