RRC ID 45911
著者 Kage-Nakadai E, Imae R, Suehiro Y, Yoshina S, Hori S, Mitani S.
タイトル A conditional knockout toolkit for Caenorhabditis elegans based on the Cre/loxP recombination.
ジャーナル PLoS One
Abstract 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.
巻・号 9(12)
ページ e114680
公開日 2014-1-1
DOI 10.1371/journal.pone.0114680
PII PONE-D-14-40651
PMID 25474529
PMC PMC4256423
MeSH Animals Animals, Genetically Modified Caenorhabditis elegans / genetics* Gene Knockout Techniques / methods* Genetic Engineering Genetic Vectors Green Fluorescent Proteins / biosynthesis Green Fluorescent Proteins / genetics Integrases / genetics Organ Specificity RNA Interference
IF 2.74
引用数 11
WOS 分野 BIOCHEMISTRY & MOLECULAR BIOLOGY
リソース情報
線虫 tm2700 tm234 tm246