cloning pcr from VENT,Pfu etc.

rogerM at qimr.edu.au rogerM at qimr.edu.au
Mon Nov 8 00:32:48 EST 1993


 Dear clonetters, 
     Does anyone have experience cloning pcr fragments from the other
   thermostable polymerases particularly VENT and DEEPVENT from NEB 
   and PfuI from Stratagene. What successes and/or failures have people
   experiences. Also has anyone had problems cloning pcr fragments
   using degenerate primers containing Inosines. We have had one
   success with these primers but other different  degenerate primers
   have been totally unsuccessful. There are white colonies showing
   up but the majority show up to be deletions if >100bp in the insert
   and unsequencable due to apparent deletions in the polylinker. The
   fragments were tested against control fragments from specific pcr
   primers by self-ligation and the test set totally failed whilst the 
   controls all self ligated.Any tips, suggestions or spelling flames
   are welcome :-). As a final note any tips to reduce my golf handicap
   from seasoned pcr golfers welcome?!!!
    Roger Mitchell.
   e-mail: rogerM at qimr.edu.au
   



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