Reverse genetics by chemical mutagenesis in Caenorhabditis elegans.
Traditional reverse genetics on yeast, mice and other organisms uses homologous recombination with transgenic DNA to interrupt a target gene. Here we report that target-selected gene inactivation can be be achieved in Caenorhabditis elegans with the use of chemical mutagens. We use PCR to selectively visualize deletions in genes of interest; the method is sensitive enough to permit detection of a single mutant among more than 15,000 wild types. A permanent frozen mutant collection of more than a million mutagenized animals has been established, and deletion mutants of several G-protein genes were isolated from it. The approach is suitable to be scaled up for systematic inactivation of all 17,000 C. elegans genes. Because it requires no transgenesis or cell culturing, it may also be applicable to small organisms usually considered to be outside the realm of reverse genetics (for example, other nematodes and insects). Any sequenced gene in any organism that can be handled in very large numbers can possibly be targeted in this way.
Jansen
G
G
Division of Molecular Biology, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
Hazendonk
E
E
Thijssen
K L
KL
Plasterk
R H
RH
D000818:Animals; D017173:Caenorhabditis elegans / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D017931:DNA Primers; D005020:Ethyl Methanesulfonate / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000235:genetics; D015723:Gene Library; D007313:Insecta; D051379:Mice; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D017384:Sequence Deletion; D014307:Trioxsalen / Q000494:pharmacology; D014466:Ultraviolet Rays
D017931:DNA Primers; D005020:Ethyl Methanesulfonate; D019204:GTP-Binding Proteins; D014307:Trioxsalen