about ligation and transformation
Posted 29 August 2003 - 05:27 AM
i never get a terrible trouble like this. it's a normal ligation and transformation. the vector is 13kb and the insert is 3kb, be used in HindIII site. i have transforme several times, and there is no any clone on plate. the efficiency is about 10-7/ug.
i have designed severy controls: the small clone vector digested by EcoRI, the self-ligation, w/wo dephosphorylation, another one is the vector added the insert as the v/i rate 1:3. at last the self-ligation is normal but when i added the insert, there is no clones.
even i transformation by electro-transformation.
i have no idea now.
anyone can help me?
thanks a lot
Posted 29 August 2003 - 09:45 AM
Serge Champetier Ph.D.
Posted 30 August 2003 - 04:22 PM
l never used the method by phenol (CHCL3?) extraction, and l don't know how to do it. but in our lab, we also use the glass wool or something like it to recove the DNA bands.
sometimes we also precipitate the ligation solution. l think l should try it.
Posted 02 September 2003 - 02:20 PM