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Point mutation in sanger sequencing - (Nov/10/2015 )

Hi guys.

I performed a sanger sequencing on both a cDNA and a DNA from the same sample. I found a point mutation in the cDNA but not in the DNA. 


Have you ever happened something like that?!

-Preziosa Buonocore-

The first thing to check is the sequencing itself.


It is not unusual for the computer to call the wrong nucleotide. Check the raw trace to see if the "mutation" in the cDNA is real before doing any other kind of troubleshooting.


Post-transcription mutation of mRNA is rare but not unknown. But I agree that first step should be looking at the trace of the sanger sequencing, and likely repeating it. PCR errors either in the initial RT-PCR or in subsequent amplification could explain this.