Q:QuickChange and deletions
hadia at bdg10.niddk.nih.gov.removethis
Wed Jan 7 22:04:58 EST 1998
I am just wondering if someone has experience in deletions (let's say 100
bp) using the Stratagene QuickChange kit. I have found that the method is
working pretty good with point mutations (at least 90% eff.) but for some
primers it needed some fiddeling around with the temperature profile to
get it to work. Any suggestions about primer design when doing deletions?
Stratagene suggests a formula to calculate the Tm where the "percentage
of mismatch" is taken into account. Do you consider "mismatches" in loop-
out mutations at all? I'd say that there is no mismatch since the primer
anneals perfectly to the template (but not the other way around...).
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