Cleaning up PCR products for BSP - Any experience with ExosapIt? (Jul/11/2007 )
I an really annoyed by the agarose clean-up of my PCR products for BSP, lately.
Has anybody here tried to use PCR purification kits or ExosapIt?
Nothing against a tan, but hours in the dark room ... ?!?
We have used ExosapIt a lot and found it works great. After PCR, I run a gel using 5 ul of PCR product. If the bands are clean and strong, just treat the remaining product with ExosapIt followed by sequencing. If there are non-specific bands besides the desired band, then gel purification is needed.
great to hear that! I'll give it a try.
The clinical staff in our lab have been using it for ages with no problems. This was for regular PCR clean ups, not BSPs, but it shouldn't make a difference. I agree its so much easier than agarose gel cut ups! We sometimes use another method called Ampure with magnetic beads, but exosapit is still easier.
I believe the advantage that Ampure offers is a product size cutoff, is that correct? So it would allow for primer dimer removal? ExoSap will only deactivate single , stranded DNA, leaving the dimers intact...
Yes I Ampure will allow for size cutoff, getting rid of primers and dNTPs. ExoSap I think also degrades the primers (the Exonuclease element of ExoSapIT) so unless the primers form terrible dimers, the ExoSapIT should get rid of these too.