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excess rna sample affect qpcr?


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#1 soymilk14

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Posted 25 September 2013 - 10:10 PM

i am just wondering if adding too much rna to the RT reaction volume can affect the qpcr later on?

 

i used 2 ug of rna template instead of the 1 ug mixed with 0.5 ul of Oligo dT( 1 ug/ul). will this affect the quality of my cdna? and is the remaining RNA affect the subsequent QPCR? thank you..



#2 pcrman

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Posted 26 September 2013 - 08:46 PM

2 ug RNA should be fine and will not affect downstream application. To detect low abundant mRNA, increasing the starting RNA is a common practice.. 



#3 jerryshelly1

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Posted 30 September 2013 - 06:01 AM

I usually use 2ug of RNA and get good, consistent results.  The amount of RNA will not affect the qPCR, because your cDNA protocol should require the degradation of your RNA products following Reverse Transcription.

 

Edit: The ending amount of RNA will not affect your qPCR data.


Edited by jerryshelly1, 30 September 2013 - 06:02 AM.





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