Hey there everybody!
I am writing because i have some doubts on an experience of my own. I am performing qRT-PCR for exiqon individual assays , and i want to get an absolute quantification of microRNAs in my samples. My doubt is: what RNA sample do i use to perform the standard curve? i am currently using reference rna from prostate tissue (Ambion) because my work is related to prostate tissues, but some of the miR's i am testing are not amplifying neither in samples, which would be plausible if they are not expressed in tumors, nor in the standards. Any tip?
Thank you,
Pissitu
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kaveh
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Posted 13 May 2012 - 03:46 PM
As long as the target microRNA is expressed at a basal level, that makes a good sample for standard curve. If the specific microRNa that you are working with is known to be a low-abundance microRNA, then you need to increase the starting material. For Exiqon, you need at least 100 copies of your microRNA to be able to detect it with SYBR green.
