how to quantify pcr products

Deanne Bell dbell at qnis.net
Wed Nov 8 12:33:57 EST 2000


Hi Karen
I found a set of markers that are designed for quantifying DNA - "Precision
Molecular Mass Standards" by BioRad  (I think the catalog # is 170-8297 but
you better check)
I use these markers to quantify my DNA preps before doing AFLPs and other
PCR techniques.  I found that the spec gave anomalous readings and I used a
lot of my DNA prep to take the A260 and A280.  After trying other methods
(DyNA Quant & DNA dipsticks) , I chose to run 3ul of my DNA prep out on an
agarose gel (0.8% - 1.2%) with these markers; stain with EtBr and then do a
spot densitometry reading using our imaging system (Alpna Innotech).  It
works great!
I have never tried it with PCR products, but I imagine that it would work
if your purified products have enough DNA to fall within the range of the
markers.
Hope this helps :-)

Deanne Bell
Molecular Markers Lab Technician
USDA Agricultural Research Service
2021 S. Peach Ave.
Fresno, CA 93727
e-mail: dbell at qnis.net
phone: 559 453-3170
fax: 559 453-3088

----------
> From: Karen Roberts <leehrat at mindspring.com>
> To: methods at hgmp.mrc.ac.uk
> Subject: how to quantify pcr products
> Date: Tuesday, November 07, 2000 3:08 PM
> 
> I have tried using spec measurements but the yield is so low after
> 
> I have purified the products that the readings give me next to or
> nothing.
> 
> Next, I have run samples on a gel and dna is there but how can I
> quantify it?  I tried comparing the bands to bands of the marker (I
> called the company and got the formula for how much dna each marker band
> has based on how much you loaded of the marker) but I am getting numbers
> that make no sense when I do this gel estimation.
> 
> I can give further details if you wish to email me privately but for
> now, what do others do to quantitate the amount of pcr product purified?
> 
> MF
> 
> 


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