Gene mutations discovery
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Entering edit mode
6.8 years ago

Hello!

I'm approaching a dna mutation analysis for the first time. I have dna sequences of a structural gene (454 reads, about 450 bp, single end) sequenced in 10 samples and I need to find mutations and quantify the abundance of each mutant in each sample. 

This is how I'm thinking to proceed:

-demultiplexing sequences using qiime

-trimming low quality bases and filtering reads too short

-clustering them at 100% similarity using usearch

-aligning centroids to some sequences found at NCBI using mummer

My questions are:

first of all, is my approach correct? Do anybody has experience in this kind of analysis and which approach used?

then, I don't know if mummer is the good choice... It is designed for snp discover in genomes, so I don't know if it could be a good idea using it for sequences that I expect to be very similar among them. If not, any other suggestion about softwares to use?

 

thanks

francesca

 

 

 

SNP mutation mummer • 1.5k views
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Entering edit mode
6.8 years ago

I'd skip clustering and just align everything. Just use a few more cores. I'd give BWA a try rather than mummer, mostly because BWA typically produces good quality results in downstream variant calling and also because it's reasonably fast.

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Hi! Thanks for your prompt reply! I wondered about using bwa or bowtie, but    using them wouldn' t imply losing new variants? I could just quantify the number of sequences matching those I have as references. Is it right?

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BWA is probably the most popular aligner used to find new variants, so no it won't lose them (unless the sequence is very highly divergent).

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