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Conditional knockout (cKO) based on site-specific recombination (SSR) technology is a powerful approach for estimating gene functions in a spatially and temporally specific manner in many model animals. In  Caenorhabditis elegans (C. elegans) , spatial- and temporal-specific gene functions have been largely determined by mosaic analyses, rescue experiments and feeding RNAi methods. To develop a systematic and stable cKO system in  C. elegans , we generated Cre recombinase expression vectors that are driven by various tissue-specific or heat-shock promoters. Validation using Cre-mediated fluorescence protein inactivation or activation systems demonstrated successful Cre-dependent  loxP  excision. We established a collection of multi-copy Cre transgenic strains for each evaluated vector. To evaluate our Cre/ loxP -based cKO system, we generated  sid-1  deletion mutants harboring floxed  sid-1  single-copy integration (SCI) using ultraviolet trimethylpsoralen (UV/TMP) methods.  sid-1  mutants that were rescued by the floxed  sid-1  SCI were then crossed with the  Pdpy-7::Cre  strain for cKO in the hypodermis. The  sid-1  cKO animals were resistant to  bli-3  RNAi, which causes the Bli-phenotyple in the hypodermis, but they were sensitive to  unc-22  RNAi, which leads to twitching of the body wall muscle. Our system, which is based on the combination of a transgenic Cre collection, pre-existing deletion mutants, and UV/TMP SCI methods, provided a systematic approach for cKO in  C. elegans .

A Conditional Knockout Toolkit for Caenorhabditis elegans Based on the Cre/loxP Recombination