XhoI problems/Footprinting

Richard P. Grant see_sig at cmtech.co.delete.uk
Tue Jun 16 07:57:14 EST 1998


In article <35866681.E0D0EC96 at uea.ac.uk>, Greg <g.dean at uea.ac.uk> wrote:

. . .

: band. Sounds straight forward. I cannot get complete digestion with XhoI
: (~50%), which is dramatically reducing my yield. I have used different
: plasmid preps, different batches of XhoI (all well within their shelf
: life), with very extended incubation times, all  to no avail. All the
: digest conditions are perfect for the enzyme, and sufficient enzyme is

Sorry to be obvious, but have you got BSA in the rxn buffer?  Spermidine
often helps too.  Try 150 µg/ml BSA and 2 mM spermidine.  

: being used. To top it all my qiagen qiaex is only working to about 60%
: recovery.

can you get the Xho digest to work /before/ gel purification?  If so, try
a different gel purification method.  We'll sell you one :-)

R

-- 
Richard P. Grant MA DPhil  | rgrant at  cmtech.co.uk
Senior R&D Scientist       | work: http://www.cmtech.co.uk/
Cambridge Molecular        | home: http://www.avnet.co.uk/adastra
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