From owner-celegans@net.bio.net Sun May 02 23:55:00 1999 Path: biosci!biosci!not-for-mail From: Michael Koelle Newsgroups: bionet.celegans Subject: new C. elegans gene knockout protocol now available Date: 2 May 1999 17:55:34 -0700 Organization: BIOSCI International Newsgroups for Molecular Biology Lines: 35 Sender: daemon@net.bio.net Approved: wmorgan@acs.wooster.edu Distribution: world Message-ID: <7gis66$gjv@net.bio.net> NNTP-Posting-Host: net.bio.net My lab has had quite good success over the past several months knocking out C. elegans genes. We use a procedure similar to that developed at NemaPharm, in which mutagenized worms are grown in microtiter dishes, stored frozen, and their DNA is screened for deletion mutations by PCR. The method requires an initial investment of ~3 weeks of work to construct the frozen mutant "library", which can be stored indefinitely and can be screened about 200 times. Once the library is constructed one can isolate a mutant in a gene of interest in only 2-3 weeks of work. We have so far succeeded in obtaining one to three mutant alleles of every gene we have attempted to knock out. We think the method works well enough that many labs may want to try it. We have prepared a very detailed protocol describing all aspects of the procedure. We hope by making this available we can minimize the difficulties other researchers might face when first trying this somewhat involved technique. The protocol can be accessed via my lab's Web site: http://www.info.med.yale.edu/mbb/koell e/ -- Michael Koelle Assistant Professor Department of Molecular Biophysics and Biochemistry Yale University School of Medicine 333 Cedar Street PO BOx 208024 New Haven, CT 06520-8024 Tel: (203) 737-5808 FAX: (203) 785-6404 .