Hi all experts,
Frist of all, please accept my apology if this question isn't relevant to here, however, I found that this forum is more active others, so please let me consult with you.
I have got Illumina sequencing reads from a healthy and diseased mouse. I want to compare the simple sequence repeats (SSR) between healthy and disease groups and discover the disease-related SSR markers. This is my first experience on this issue, so I was wondering what is the suitable pipeline for this work? any comment and suggestion would be highly appreciated.
Thanks in advance
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Hello,
Ok so the DE analysis seems good and this would give you an idea of some genes that might be involved in the disease in your mice. The fact that they're not isogenic does mean that some DE could be down to random differences between the mice though. But if you get a handful of DE genes you can look at putative functions and pathways that might be involved. You don't need to do multiple assemblies, just one, and you can assemble all your data at once then map the reads individually.
From this you'll do DE analysis and you can look at alignments of specific genes to look for seq differences between WT and diseased mice. So if there's an SSR (maybe 5 or 10% of genes have SSRs?) then you might find differences (again not all SSRs are polymorphic). So I don't see why you have to find an SSR and it is unlikely you'll find a polymorphic SSR that differentiates your WT and diseased mice.
So whats the need for this SSR? What are you trying to do with it? Do you want a marker that differentiated between WT and diseased mice? IMHO I find this unlikely to work. But there might be other things you can do.
Cheers, Mark
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Hi, Sorry if I didn't answer that part. Once you map the reads to the de novo you'll output a bam file for each individual. This can then be converted to vcf and fasta files using samtools. You can then align the transcripts and check for polymorphism between diseased and susceptible.
Best wishes, Mark
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Hi, Assembling a transcriptome using a published genome, instead of de novo, should reduce the assembly of spurious transcripts. In your case if you're looking for an SSR you will be able to scan the genome near your candidate gene (eg a DE one) and looks for an SSR motif.
Best wishes, Mark
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