ss. DNA restriction (yes, they cut)

Wed Oct 5 16:04:53 EST 1994

William Bourn writes:

> I have been trying to find out if restriction enzymes of the type that are
> commonly used for cloning etc have any activity on single stranded DNA. 

This was a topic of investigation in the mid 1970's.  Initially several
R. enzymes were found to cut "single-stranded" phiX 174 by Robert Blakesley
working with Robert D. Wells.  That group interpreted their results in terms
of local ds. str.  Others (I'm sorry I don't remember who) thought the enzymes
directly cut true single strands and showed the cutting of synthetic oligo
nucleotides.  However, since the R. sites were palindromic one couldn't rule
out some residual tendency of the oligos to base pair.  Subsequently, I
remember someone cleanly cutting Hinf fragments out of single stranded
DNA.  Hinf has a non-palindromic sequence [GANTC].  I think the resolution
of the issue was that most if not all R. enzs. would directly cut single
stranded DNA but a very low rates (<1% the ds. rate).  Some enzymes could
handle the short ds. regions in "single stranded" molecules like phi X
+ strand and cut them at higher rates (but still < the ds. rate).  Here is
an non-exhaustive ref. list:
Blakesley et al., JBC 252:7300-6 (1977)
Been and Champoux, Meth. in Enzymol. 101:90-8 (1983)
Wells and Neuendorf, Gene Amplif Anal. 1:101-11 (1981)

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