Deletion Problems using recombinant PCR

D L Clarke mb935645 at silver.shef.ac.uk
Wed Feb 7 06:18:21 EST 1996


Dear colleagues,

I would like to create a  deletion in gene using recombinant
 PCR.  I have previously created a restriction site by changing
two base pairs.  However, I have been unsuccessful in creating
a deletion.  The two primary PCR products can be created and have been
verified.  But for the final PCR reaction with the two outside primers
I cannot obtain a product.  The two internal primers are 30bp in
length this should be enough to anneal and allow the second
reaction to occur.

I would be most grateful if anybody has any suggestions on the conditions
required to produce the deletion.

Many thanks

Dave Clarke

D.L.Clarke at Sheffield.ac.uk






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