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Nanodrop vs Picogreen to quantify post-PCR product

pcr purification product real-time pcr nanodrop picogreen

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#1 detriar

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Posted 03 March 2014 - 07:29 PM

Dear all,

 

I wonder if it is the right subforum to ask, but since it has something to do with PCR product, I'm gonna ask here.

 

I purified my PCR product (excised the single band, and using column from roche kit to purify it) and I want to use this product as standard for the absolute quantification real-time PCR project. Thing is, I have to know the exact concentration of this purified PCR product, so that I can convert the concentration (ng/ul) to copy molecules/ul using Avogadro equation. From the journal, the researchers use picogreen to quantify that purified PCR product, but since I don't have such the equipment nor the reagen, I wonder if it is okay to quantify the purification product's concentration with Nanodrop?

 

I really hope you guys can help me with my question.

 

Thank you.

 

Cheers,

 

Dee



#2 phage434

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Posted 03 March 2014 - 07:52 PM

It's probably ok to use the Nanodrop after purification. Picogreen is good because it is less sensitive to ssDNA or nucleotides.  But you have eliminated most of those with gel purification.



#3 hobglobin

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Posted 04 March 2014 - 08:27 AM

You have to consider that you also lose DNA during the purification.


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#4 detriar

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Posted 23 April 2014 - 02:10 AM

Thank you :)







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