Direct PCR from E. coli colonies

Steven Sullivan sullivan at gwis2.circ.gwu.edu
Tue Jul 11 15:57:10 EST 1995


I touch a 10ul pipette tip to the colony and stick it in a 25 ul standard
PCR reaction with 1ul each of T7 and SP6 primer (Promega), swirl it, do
oil overlay, and cycle at 94,5' //94,30";  52,30"; 72,60" (30 sec for
inserts <1 kb) x 30 //72, 5'. I've PCRd up pieces ranging between 0.2 and
2.5 kb from Bluescript and TA vectors with no problem.  I use thin-wall 
tubes and a Barnstead Thermolyne cycler. 

I use pipette tips, as I said; I read recently in Biotechniques that wooden 
toothpicks may have their own contaminating or inhibitory effect, and 
should be avoided.  



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