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Question about miRNA-star and Taqman Q-PCR miRNA array validation


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#1 rnarm

rnarm

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Posted 17 November 2009 - 09:54 AM

Hi. I have 2 questions on miRNAs I was hoping someone here might be able to answer. Firstly do miR-star strands target differential mRNAs to the non-star strand. If so, are these targets accounted for in prediction databases e.g. TargetScan pulls miR family targets.

Secondly, I have conducted a few miRNA Q-PCR arrays which I am currently validating. The problem I have is the RT primers for the array are cyclic (40 cycles) but single assays consist of using a single non-cyclic RT reaction. This causes my endogenous control (u6 or RNU48) to come up much later in the cycles compared to the arrays. So my question is, if my endogenous is CT 26 for U6 in the single assays, can anyone here comment on what CT would be moderately expressed miRNA? I ask this because miRNAs I initially selected from my arrays that were coming up at CT 30 (U6= CT19) are coming up too late for single assay validation (CT 37).


Thanks in advance,

Regards,

Richard

:)




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