I have desinged and tested primers for RT-PCR, then purified the PCR products from the gels and send them to sequence at GATC (SUPREMERUN) using the forward primers.
After blasting the reads to refseq, I see that they are ~30% shorter than the PCR product. Example:
PCR expected length = 182 bp Read length = 133 bp
I must mention that in the gels the expected PCR sizes were ok.
Does anyone have an idea what is the reason for this decrease in length?