Does anyone know how...

Tom Knight tk at csail.mit.edu
Thu Nov 4 16:46:58 EST 2004


"cloningman" <xxxgifalone at tiscali.it> writes:
> ...to ligate an ssDNA oligo of 15-20 nt to a longer ssDNA sequence (obtained
> from an asymmetric PCR)? Which enzyme does that?
> Thanks to all in advance

T4 RNA (not DNA!!) ligase will ligate most anything to anything else.
You still need the 5' phosphate, so make sure that it is present on
the 5' end of the fragment on the 3' end of your desired result.  You
want to make sure it is *not* present at the 5' end of the fragment at
the 5' end of your final result, to avoid unwanted side-reactions.
Even so, circularization of your phophorylated strand will occur.  You
can minimize this with an excess of the nonphosporylated strand.






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