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Alternative splicing or non-optimal PCR conditions?

Alternative splicing or non-optimal PCR conditions? - RNA Techniques Forum

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Old 01-24-2013, 07:34 PM
Pipette Filler
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Default Alternative splicing or non-optimal PCR conditions?



Hello,

I have performed a PCR on genomic DNA and after electrophoresis I obtained just one band. However, when I performed a PCR on cDNA (after retrotranscription) with the same primers I obtained 2 or three bands, depending on the sample. How can I know if such different bands result from "alternative splicing" of the mRNA or if it is just an artifact due to non-optimal PCR conditions? Is it possible to clone these PCR products (from cDNA) even if there is no a single band? I have tried to sequence after extracting each band from the gel, but not enought DNA...

Thanks in advance for your help!
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Old 01-25-2013, 04:39 PM
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Default Re: Alternative splicing or non-optimal PCR conditions?

You could try a Northern blot to see if you also get the three bands -- that would eliminate the possibility that the three bands are due to a PCR artifact.
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