From owner-dros@hgmp.mrc.ac.uk Thu Oct 17 22:59:32 2002 Return-Path: Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with SMTP id 84F6C17B33 for ; Thu, 17 Oct 2002 22:59:29 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 32AFA17A86 for ; Thu, 17 Oct 2002 22:59:27 +0100 (BST) Received: by mercury.hgmp.mrc.ac.uk (Postfix, from userid 6020) id 6705517B2E; Thu, 17 Oct 2002 22:59:25 +0100 (BST) Received: from localhost (localhost [127.0.0.1]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id 590AA17AA6 for ; Thu, 17 Oct 2002 17:47:49 +0100 (BST) Received: from dfw7-1.relay.mail.uu.net (dfw7-1.relay.mail.uu.net [199.171.54.106]) by mercury.hgmp.mrc.ac.uk (Postfix) with ESMTP id B666917ADD for ; Thu, 17 Oct 2002 17:47:30 +0100 (BST) Received: from itssrv1.ucsf.edu by dfw7sosrv11.alter.net with ESMTP (peer crosschecked as: itssrv1.ucsf.edu [128.218.95.22]) id QQnkzj14973 for ; Thu, 17 Oct 2002 16:47:29 GMT Received: (from news@localhost) by itssrv1.ucsf.edu (8.11.3/CDR-8.11.3) id g9HGlSo40310; Thu, 17 Oct 2002 09:47:28 -0700 To: bionet-drosophila@uunet.uu.net From: Jean-Karim Newsgroups: bionet.drosophila Subject: Re: Schneider S2 cell culture Organization: UCSF, ITS Message-ID: <3DAEEAC7.7090201@cgl.ucsf.edu> References: <5.1.0.14.0.20021017150326.026350f0@pop.toulouse.inserm.fr> Mime-Version: 1.0 Content-Type: text/plain; charset=ISO-8859-1; format=flowed Content-Transfer-Encoding: 8bit User-Agent: Mozilla/5.0 (Macintosh; U; PPC; en-US; rv:1.0.1) Gecko/20020823 Netscape/7.0 X-Accept-Language: en-us, en Date: Thu, 17 Oct 2002 22:59:25 +0100 (BST) X-Razor-id: 839dfc51889d36eda3774c30503bd74a35214643 Sender: owner-dros@hgmp.mrc.ac.uk Precedence: bulk Bonjour Pour les cellules S2, nous utilisons le milieu de Schneider avec 10% de serum de veau inactive a la chaleur. Les cellules adherent peu au plastique et se decrochent en pipettant. Les amas peuvent se separer en pipettant plusieurs fois. Le comptage se fait avec un hemacytometre. Generalement, on ensemence a 1-2.10^6/ml et on dilue environ au 1/5-1/10 tous les trois jours. Pour ce qui est des transfections, la coprecipitation au phosphate de calcium marche mais le rendement est faible (5-10% des cellules sont transfectees). Avec le Fugene6 de Roche, j'arrive a 40-50% de cellules transfectees. Pour cela, j'ensemence les cellules a 2.10^6/ml et je suit le protocole en utilisant un rapport de 3µg d'ADN pour 3µl de reactif. Bonne chance. Jean-Karim Jean-Karim Hériché Dpt. of Biochemistry and Biophysics University of California, San Francisco Room HSE 901 513 Parnassus street San Francisco, CA 94143-0448 U.S.A. Phone: 415-476-4709 .