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Real Time PCR Reaction minimum gDNA required

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Real Time PCR Reaction minimum gDNA required

Postby talt1412 » Mon Nov 02, 2009 9:56 pm

Hi,
I would like to do Real Time PCR reaction with genomic DNA (Genotyping by TaqMan) I have only Plasma samples and I have tried to extract DNA from them and received very low concentration of 3ng/ul (measuring by Nanodrop), is this concentration enough for the Real Time PCR reaction? I know it can measure from 0.1ng of genomic DNA but I am not sure about my concentration since it has been measured in Nanodrop which is not so accurate ...
thanks in advance for any help in this issue
tal
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Postby JackBean » Tue Nov 03, 2009 7:25 am

Question is, how many copies should you have with such amount. As you write about plasma samples, you isolate from human?

What about to isolate from larger amount and than put it together? ;)
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Postby canalon » Tue Nov 03, 2009 4:50 pm

Start from other a better/easier to extract source of DNA. Prepare a standard which will cover higher and lower concentration of DNA. Run in triplicate, check the minimal concentration to get a reliable and repeatable signal, this will tell you the minimum amount of DNA that can be used for your particular set of primer/probe/condition/PCR mix.
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