5' filling with Klenow
mkennedy at chmeds.ac.nz
Mon Sep 12 19:59:48 EST 1994
In article <1994Sep13.124605.594 at chmeds.ac.nz>, mkennedy at chmeds.ac.nz (Martin Kennedy) writes:
> In article <35289a$te at mark.ucdavis.edu>, ez003450 at chip.ucdavis.edu () writes:
> I would definitely sequence the junction. Don't forget that Klenow still
> has a 3'->5' nuclease activity, and it can remove part of the 5' overhang.
On second thoughts this isn't a sensible explanation for the occasional
losses of bases seen at these sites. Perhaps it is caused by 5'->3'
nuclease present as a contaminant in some component (DNA prep, enzyme etc)
of either the digestion or fill-in reaction. Whatever the cause, loss
of bases can occasionaly occur after these sorts of manipulations, and it
would be wise to sequence.
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