Sanger sequencing chromatogram aligned to reference gene

Question

[vujex]

I did Sanger Sequencing on 10 samples targeting one specific gene to see whether we detect a particular SNP. We did forward, reversed and tested three primer sets so there are quite a lot samples in total.

Now, I am wondering if there is a quick way to generate chromatograms of the data so for all the 10 samples, aligned to the reference genome? This would save a lot of time compared to the conventional way (aligning via Blast ect.).

[Asaf]

We Used to do it using sequencher. It is manual and laborious but we went through hundreds of chromatograms to verify the sequencing.

[Joe]

You can do this in Snapgene pretty easily. I'm not aware of a commandline tool for something like this however.

Answer 1

[GenoMax]

I am wondering if there is a quick way to generate chromatograms of the data so for all the 10 samples, aligned to the reference genome?

I assume you already have the chrromatograms having sequenced the samples using sanger otherwise there is no way to generate the chromatograms.

There are two programs (novoSNP and Tracy) mentioned in the answers for this past question (A: Sanger sequencing data analysis ) that should work with your sequence files. Both are free compared to commercial applications.

Answer 2

[trausch]

Indeed, you can use tracy as a command-line application to process them in batch or use the online web applications:

• Trace alignment: https://www.gear-genomics.com/sage/
• Variant calling: https://www.gear-genomics.com/indigo/
• Multiple trace alignment (reference-guided or de novo): https://www.gear-genomics.com/pearl/