High-throughput screening of small molecules for bioactivity and target identification in Caenorhabditis elegans

被引:84
作者
Burns, Andrew R.
Kwok, Trevor C. Y.
Howard, Al
Houston, Ed
Johanson, Karl
Chan, Anthony
Cutler, Sean R.
McCourt, Peter
Roy, Peter J.
机构
[1] Univ Toronto, Terrence Donnelly Ctr Cellular & Biomol Res, Dept Med Genet & Microbiol, Toronto, ON M5S 1A8, Canada
[2] Univ Toronto, Dept Cell & Syst Biol, Toronto, ON M5S 1A8, Canada
[3] Univ Toronto, Collaborat Program Dev Biol, Toronto, ON M5S 1A8, Canada
[4] Elegenics Inc, Mountain View, CA 94043 USA
基金
加拿大健康研究院; 加拿大自然科学与工程研究理事会;
关键词
D O I
10.1038/nprot.2006.283
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
This protocol describes a procedure for screening small molecules for bioactivity and a genetic approach to target identification using the nematode Caenorhabditis elegans as a model system. Libraries of small molecules are screened in 24-well plates that contain a solid agar substrate. On top of the agar mixture, one small-molecule species is deposited into each well, along with worm food (E. coli), and two third-stage or fourth-stage larval worms using a COPAS (Complex Object Parametric Analyzer and Sorter) Biosort. Three to five days later the plates are screened for phenotype. Images of the wells are acquired and archived using a HiDI 2100 automated imaging system (Elegenics). Up to 2,400 chemicals can be screened per week. To identify the predicted protein target of a bioactive molecule, wild-type worms are mutagenized using ethylmethanesulfonate (EMS). Progeny are screened for individuals resistant to the molecules effects. The candidate mutant target that confers resistance is then identified. Target identification might take months.
引用
收藏
页码:1906 / 1914
页数:9
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