.ab1 exoms analysis

[_y_s]

Hi all,

I'm new in this site and also not familiar with what I'm working with, so excuse me if the answer exist in another thread.

Actually, I have many exons sequences on a gene, that I want to analyze to find eventually some mutations (SNPs, CNVs indels). The exons are sequenced using capillary sequencing (Sanger) and are in .ab1 and .seq formats.

So I'm looking for any advices and/or tutorials to deal with.

thank you so much

.Y.S.

[sklages]

Hi all,

I'm new in this site and also not familiar with what I'm working with, so excuse me if the answer exist in another thread.

Actually, I have many exons sequences on a gene, that I want to analyze to find eventually some mutations (SNPs, CNVs indels). The exons are sequenced using capillary sequencing (Sanger) and are in .ab1 and .seq formats.

So I'm looking for any advices and/or tutorials to deal with.

thank you so much

.Y.S.

Preprocess your ab1 data with something like 'lucy' [1] or 'Lucy2'[1a].

For mapping and SNP calling you may find various solutions,

- PolyPhred (non-commercial) [2]
- GeneScreen (non-comemrcial) [3]
- Genious Pro [4]
- Mutation Surveyor [5]

This list is far from being complete; have a look at the software hub of seqanswers and/or ask Google.

hth, Sven

[1] = http://lucy.sourceforge.net/
[1a] = http://www.complex.iastate.edu/downl...cy2/index.html
[2] = http://droog.gs.washington.edu/polyphred/
[3] = http://dna.leeds.ac.uk/genescreen
[4] = http://www.geneious.com/web/geneious/geneious-pro
[5] = http://www.softgenetics.com/mutationSurveyor.html

[maubp]

Preprocess your ab1 data with something like 'lucy' [1] or 'Lucy2'[1a].

You could simply convert from the ab1 files direct to FASTA+QAUL or FASTQ using something like EMBOSS seqret or Biopython - but this would give you the base calls inside the abi file, typically made on the original instrument. This is where Lucy or tracetuner or similar comes in - they re-analyse the chromatogram traces and should give a better set of base calls than the defaults.

[sklages]

You could simply convert from the ab1 files direct to FASTA+QAUL or FASTQ using something like EMBOSS seqret or Biopython - but this would give you the base calls inside the abi file, typically made on the original instrument. This is where Lucy or tracetuner or similar comes in - they re-analyse the chromatogram traces and should give a better set of base calls than the defaults.

Oh, yes, you are right. I forgot to mention that it is a good idea to re-basecall the ab1 files with 'phred' or 'tracetuner'. But all that basecalling / preprocessing stuff is only necessary when you are going to use polyphred as all these commercial packages are feed directly with ab1 files and all the preprocessing can be performed within the software itself.

hth, Sven

[_y_s]

Thanks to both of you very much, I'll follow your instructions and I'll let you know if I've got some issues.

Thanks again

YS