QRT-PCR - (Apr/11/2005 )

Do you have any idea, that the I have clearly specific melting curve peaks , also I can see the specific amplification on the gel. There is no quantification value for samples, they give very bad amplification curves when they analysed.

Also my specific gene has got at least 9 alternative splicing variants, and I am trying to amplification without a specific prob. My pirmers are designed to shared-common part of the cDNA. Should I use a specific prob, is it sorce of the problem?

Thanks

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Hi,

What do you mean by bad amplification curve? You don't see the three stages or the delta R values are too small? Are you using random hexamers or oligo dt to synthesize the first strand?

SV

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Hi

I mean I have got a specific amplification and they are my target. Bu I can not see a quantification value even for the standarts, positive controls and target samples.

And yes I used random hexamer.

SYT

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Could it be that your fluorescent dye in your mix has gone off?

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or perhaps you have an instrument problem...something wrong with the detector...or a recalibration of the spectra/dye/fluorescent values is in order.

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