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qPCR


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3 replies to this topic

#1 thegene

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Posted 27 April 2009 - 06:50 AM

Hi mates,

i am trying to do standard curves for a couple of primer pairs that i want to use for measuring the expression levels for a gene (MRNA) after mammalian cells transfection with a plasmid containing the cDNA of the gene of interest.

The standard template with known concentration that i have used for standard curves, is the original plasmid used for transfection with different serial dilutions. Is that correct??

#2 aimikins

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Posted 27 April 2009 - 08:03 AM

yeah, sure. that's the easiest way. just make sure the dilution range is nice and wide.
"it is a miracle that curiosity survives formal education" -A.E.

#3 thegene

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Posted 28 April 2009 - 12:06 AM

yeah, sure. that's the easiest way. just make sure the dilution range is nice and wide.


Thanks Aimikins,
I have performed two times and in every time i didn't get amplification plots in the qpcr, i feel that, it is not right to use plasmid DNA as a template for qPCR. Do you any idea about that?

Regards

#4 aimikins

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Posted 29 April 2009 - 01:48 PM

what is your dilution range?

also, can you add a pic of your melting curves?
"it is a miracle that curiosity survives formal education" -A.E.




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