gravatar for Rogerio Ribeiro

3 hours ago by

Hi biostars.

I have a question regarding minimap2.
For my biological problem, I am doing differential expression analysis and differential transcript usage for six (3 vs 3) samples, which have been sequenced using a cDNA protocol with MinIOn.
To start I built a genome-guided transcriptome using minimap2 + stringtie. During the alignment step this is the comand I have used:

minimap2 -t 8 -ax splice cro_v2_asm.mmi 01_filtering/after_trim/all_reads_nano.fastq > 02_annotation/all_reads.sam
samtools view -q 40 -b -F 2304 02_annotation/all_reads.sam | samtools sort [email protected] 8 -o all_reads_filtered.bam
samtools index all_reads_filtered.bam

Recently I saw a post suggesting that aligning Nanopore reads should be performed using the -ax map-ont option. However, since reading the manual, I assumed this option was reserved for genomic reads, which I assumed was reads obtained from genome sequencing.

From the minimap2 manual:

./minimap2 -ax map-ont ref.fa ont.fq.gz > aln.sam         # Oxford Nanopore genomic reads
./minimap2 -ax splice ref.fa rna-reads.fa > aln.sam       # spliced long reads (strand unknown)

The question arose as to when I was messing with GitHub pipelines for DEG, one of the pipelines used the minimap2 to map reads to a transcriptome, hence my confusion regarding the manual.

What is usually the best present when mapping ONT RNA-seq reads to a genome?

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