Targeted engineering of the Caenorhabditis elegans genome following Mos1-triggered chromosomal breaks

• Published in: The EMBO journal 2007-01, Vol.26 (1), p.170-183
• Main Authors: Robert, Valérie, Bessereau, Jean-Louis
• Format: Article
• Language: English
• Subjects: Animals; Base Sequence; Caenorhabditis elegans; Caenorhabditis elegans - genetics; Caenorhabditis elegans - physiology; Chromosomes; Computational Biology; DNA Breaks, Double-Stranded; DNA damage; DNA repair; DNA-Binding Proteins - genetics; Drosophila; Drosophila melanogaster - metabolism; DSB repair; EMBO13; EMBO43; Genetic Techniques; Genome; Genomics; homologous recombination; Models, Genetic; Molecular Sequence Data; Mutagenesis; Mutation; Nematodes; NHEJ; Plasmids - metabolism; Recombination, Genetic; targeted mutagenesis; Transgenes; Transposases - genetics
• ISSN: 0261-4189; 1460-2075
• DOI: 10.1038/sj.emboj.7601463

The Drosophila element Mos1 is a class II transposon, which moves by a ‘cut‐and‐paste’ mechanism and can be experimentally mobilized in the Caenorhabditis elegans germ line. Here, we triggered the excision of identified Mos1 insertions to create chromosomal breaks at given sites and further manipulate the broken loci. Double‐strand break (DSB) repair could be achieved by gene conversion using a transgene containing sequences homologous to the broken chromosomal region as a repair template. Consequently, mutations engineered in the transgene could be copied to a specific locus at high frequency. This pathway was further characterized to develop an efficient tool—called Mos TIC—to manipulate the C. elegans genome. Analysis of DSB repair during Mos TIC experiments demonstrated that DSBs could also be sealed by end‐joining in the germ line, independently from the evolutionarily conserved Ku80 and ligase IV factors. In conjunction with a publicly available Mos1 insertion library currently being generated, Mos TIC will provide a general tool to customize the C. elegans genome.