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-   -   Alternative splicing or non-optimal PCR conditions? (http://www.molecularstation.com/forum/rna-techniques-forum/87411-alternative-splicing-non-optimal-pcr-conditions.html)

Montojo 01-24-2013 07:34 PM

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!

Jon Moulton 01-25-2013 04:39 PM

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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