SOS!--PCR restriction cut

[Hector Cruz-Lopez]

In article <4ihofq$kvp at ccshst05.cs.uoguelph.ca> wyu at uoguelph.ca (Wenjin Yu) writes:
>From: wyu at uoguelph.ca (Wenjin Yu)
>Subject: SOS!--PCR restriction cut
>Date: 17 Mar 1996 19:17:14 GMT

>        I have a big problem to cut PCR product in which the restriction site
>is generated in the primer.  At first I designed a primer containing a Bam 
>HI site.  Based on the inforamtion in New England Biolabs Catalog, 4 
>extra bases was added at the 5'-end, but the PCR product couldn't be cut 
>no matter how much enzyme was used.  Then I made another primer 
>containing an EcoRI site, and 15 extra-bases was added to the 5'-end this 
>time.  But it seems still not work!  I was wondering if anyone can help me to
>deal with this big trouble.  Many thanks.
> (please send your HELP to my mail box: wyu at uoguelph.ca)
>Wenjin Yu
>U of Guelph


I'd recommend you try another enzyme supplier.  I have had problems digesting 
PCR products with NEB enzymes.  I switched to Boehringher Mannheim and that 
took care of my problems.  This only happened with PCR products.


Hector "Che" Cruz-Lopez
Florida Game and Fish Commission
Boca Raton, Florida