Ligation, and Transformation iNHIBITORS.

Jee-Yeong Jeong jeong at biochem.purdue.edu
Fri Mar 12 18:51:51 EST 1999


Somebody in our Lab once had the similar problem and he showed clearly
there was an inhibitory factor in his ligation reaction by comparing
some control experiments such as plasmid in water, plasmid in 1X
ligation buffer, etc. The circular plasmid in water was well transformed
into E.coli, but one in ligation buffer (I don't remember exactly
whether ligase was also present.) was not. He solved this problem by PCI
extraction and EtOH ppt after ligation reaction. I think this is not a
general problem because other batch of ligase and ligation buffer do not
usually have such a problem. So, you could try a new ligase and buffer
or PCI/EtOH like him.

amherrera at CIGB.EDU.CU wrote:

>     Hi all
>
>     I having a low transformation frequency when transform my
>     ligation?
>
>     I have evidence about the good efficiency of the competent
>     cells.
>
>     The vector I used is 10.kb and the band over 3.3kb
>
>     Thanking Any Suggestion
>
>     D.Q.V

--
===================================================
Jee-Yeong Jeong
Graduate Research Assistant
Department of Biochemistry
Purdue University
Hansen Life Science Research Building, Room 225
West Lafayette, IN 47907
Tel (765) 494-6591          Fax (765) 494-7897
Email: jeong at biochem.purdue.edu





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