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a-slide / NanoCount

EM based transcript abundance from nanopore reads mapped to a transcriptome with minimap2
https://a-slide.github.io/NanoCount/
MIT License
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Different options when using minimap #14

Closed Tomcxf closed 1 year ago

Tomcxf commented 1 year ago

I used to do the following command to map my direct RNA data by nanopore. minimap2 -ax map-ont -splice -uf -k14 the_reference_transcriptome.fa basecall_by_guppy.fastq > minimap_transcriptome.sam And when I tried the command you recommend minimap2 -t 4 -ax map-ont -p 0 -N 10 transcriptome.fa.gz reads.fastq.gz I got a larger file. Both the bam file and count file created by nanocount. I noticed you said the -N is about the secondary mapping. But when I used samtools flagstat, I found both of them have no secondary mapping. And the recommend command make more mapping. So I don't know what cause that. Should I use the recommend command to do the analysis instead of the formed command? Because the former command is recommened by Minimap offically. Thank you!

Tomcxf commented 1 year ago

oh I found the secondary you recommended is larger than former. It also cause larger mapping rate. Is that the control is less strict? Will it effect the consequence?

josiegleeson commented 1 year ago

Hello, apologies for the delayed response.

Running minimap2 with the -N 10 flag should retain up to 10 secondary alignments. You can use this command or just the default minimap2 command 'minimap2 -ax map-ont transcriptome.fa reads.fastq' and this should work fine as well if you're having issues.

There is no need to include the -splice flag when you map to the transcriptome. Only use this for the genome mapping.

Let me know if you need more help. Josie.