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differentiating ssDNA and dsDNA

ed_taboada at hotmail.com ed_taboada at hotmail.com
Fri Jan 14 12:15:43 EST 2000


Hi everyone,

I'll be doing asymmetric PCR to generate a single stranded product which
we will want to quantify. My boss worries about the amount of double
stranded product that will be created and would like to quantify the
amount of each species that will be generated by the PCR. We need to
avoid double stranded product like the plague and are even considering
cleaning-up the product by using a biotinylated primer/streptavidin
magnetic beads/denaturation. We want to start with a primarily ss
product (hence the asymmetric PCR) in order to minimize the costs
associated with the (outrageously expensive) magnetic beads.

What sort of primer ratios might allow significant generation of ss
product while minimizing the relative amount of ds product ?

Also, is there a way for us to quantify the amounts of ssDNA and dsDNA ?

If anyone could point me in the right direction I would be most
grateful.

thanks in advance,

Ed


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