Annotation error in the results generated sanger sequencer

[Scarlet-001]

The sequencer shows annotation for insertion one bp ahead instead of showing the position before and after the insertion. What could be the possible issue or is that an acceptable annotation?

[tobiasrausch]

I am not sure what the problem is? Can you please elaborate?

[ghost]

I am not sure what the problem is? Can you please elaborate?

I am not an expert in the field and was just doing my internship at a Cancer lab. I will try my best to explain.

Working with sanger is new to me. While analyzing a report we found an insertion that has not been reported in any databases so far, we were working on checking if the insertion is pathogenic or not but when we reanalyzed the sanger annotation through variant reporter of the insertion it was giving it one position ahead and we analyzed that with the VEP as well that is were the confusion started.

The sequencer showed annotation of an insertion not between the bases but a step/position next to it:

c.5532-5533insGAA is the insertion position came after position 32 and before 33 but the sequencer annotates it like c.5533-5534insGAA.

We were working on human genome hg38.

The reason to suspect the error is because considering the error it shows insertion of a codon, between a codon while my instructor suspect that may not be the case considering patient's history. My instructor believed that the insertion point is before the codon not inbetween the codon itself.

And the annotation is in this form [c.33-34insAUU]+[=] and I don't know what the +[=] represents here.

We were confused if it was the error by sequencer or if we didn't know something about the annotations.

[tobiasrausch]

Usually it's best to then look at the raw chromatogram trace signal. In Indigo the variants are linked to the trace signal plot so you can jump to the position of the variant in the trace.

[ghost]

Usually it's best to then look at the raw chromatogram trace signal. In Indigo the variants are linked to the trace signal plot so you can jump to the position of the variant in the trace.

So should I try using that software? What file should I give as an input? Or should I put the sequence?