1. Single-copy insertion of transgenes in Caenorhabditis elegans
- Author
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Chris E. Hopkins, M. Wayne Davis, Erik M. Jorgensen, Christian Frøkjær-Jensen, Blake J. Newman, Jason M. Thummel, Søren-Peter Olesen, and Morten Grunnet
- Subjects
Transposable element ,Genetics ,Mutation ,biology ,Extrachromosomal array ,biology.organism_classification ,medicine.disease_cause ,Noncoding DNA ,Germline ,chemistry.chemical_compound ,chemistry ,Extrachromosomal DNA ,medicine ,Caenorhabditis elegans ,DNA - 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.
- Published
- 2008
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