Thanks, that's really helpful to know! I may have to invest in some high volume filters early on in that case. All the best, Cathal
--- Twitter: @onetruecathal Sent from my beloved Android phone. On 25 Jul 2010 03:23, "DK" <[email protected]> wrote: In article <[email protected]>, Cathal Garvey <[email protected]... Simple calcium chloride cells are super easy to make but their transformation efficiency tops at 10^6/ug (usually an order less). There are many protocols that give higher efficiencies for CaCl2 cells but they seem to be poorly reproducioble. The most robust of all chemical protocols is Inoue method, whuch us great. Gives around 10^8/ug in our hands (don't think we ever hit 10^9 stated in the paper). Its only downside is that the frozen cells seem to slowly lose transformation efficiency over time. And there are some applications where higher efficiency with electroporation matters. Google +Inoue +transfrmation +E.coli. Growing cells at low temp is a critical part of Iniou method, so if you don't have refrigerated chaker, wrap flasks in towels and spray water on them to keep cells cool. >Has anyone got a straightforward protocol for chemically competent cells to >add? One which might ... I can't be affirmative here but my sense is that it matters to an extent to all conventional methods. DK _______________________________________________ Methods mailing list [email protected] http://www.... _______________________________________________ Methods mailing list [email protected] http://www.bio.net/biomail/listinfo/methods
