Question: Including failed minION reads in alignment
gravatar for eayasi
2.2 years ago by
eayasi0 wrote:

I am trying to get data from a minION run I have done. Most of my reads a pretty low quality (rapid seq kit with shortened fragmentation on 8kb plasmid...might be the problem?) and over half failed. enter image description here I've tried to use a few aligners like bwa-mem, graphmap, last - but nothing is aligning to my plasmid. Is it worth it to to try the failed reads? Will I get usable/ok data with enough coverage?

Is there a way to extract fastq/a files from just the failed reads using poretools or nanoOK? Any tips would be appreciated!

minion alignment • 1.0k views
ADD COMMENTlink modified 2.2 years ago • written 2.2 years ago by eayasi0

If nothing is aligning, you might try BLAST on a few hundred reads (or, in fact, BBSketch on all of them) to see if you have contamination. Is this plasmid likely to be on NCBI (RefSeq/nt)?

I'm not sure what "failed" means to Nanopore software, but it's likely that you can't extract them as fasta/fastq because the bases simply have not been called. You can't align the raw signal.

ADD REPLYlink modified 2.2 years ago • written 2.2 years ago by Brian Bushnell16k

Thanks for your reply. I have had one read align in graphmap. :/ I have blasted some and gotten yeast mitochondrial DNA (makes sense since I isolated plasmid from yeast). Might have to rethink seq protocol to get usable data.

ADD REPLYlink written 2.2 years ago by eayasi0

Did you use bwa-mem -x ont2d?

The majority of your reads are also rather short, it seems.

ADD REPLYlink written 2.2 years ago by WouterDeCoster40k
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