Nanopolish output: no alignments

[tait123]

Hello, I’m working on running nanopolish for methylation calling. My command line was: nanopolish call-methylation -t 8 -r /home/choilab_tk/nanopore/data/221108_bank7492_NIID_Cas9/20221108_0927_X5_FAV43835_7491c0d0/fastq_pass/pass/merged/output_merged.fastq -b /home/choilab_tk/nanopore/results/221108_bank7492/minimap2/merged/output_merged.bam -g /home/choilab_tk/hg38/hg38.fa -w "chr1:149,390,100-149,391,900" > methylation_calls.tsv

However, when I run above command line, output shows: [bam process] iterating over region: chr1:149,390,100-149,391,900 [post-run summary] total reads: 26, unparseable: 0, qc fail: 0, could not calibrate: 0, no alignment: 13, bad fast5: 0

Any recommendation on why 50% of total reads cannot be aligned? and how to solve this problem?

For you information, I used Guppy for basecalling, and it command line was: guppy_basecaller -i /home/choilab_tk/nanopore/data/221108_bank7492_NIID_Cas9/20221108_0927_X5_FAV43835_7491c0d0/fast5_pass -s /home/choilab_tk/nanopore/data/221108_bank7492_NIID_Cas9/20221108_0927_X5_FAV43835_7491c0d0/fastq_pass -c dna_r10.4.1_e8.2_260bps_fast.cfg --num_callers 4 --trim_adapters --trim_primers --compress_fastq