probability_modified column is empty

[BioinfoHub-PeiQinNg]

Hi there,

I have tried running m6anet run-inference to predict m6A modifications on my nanopore data.

This was done based on the quick start tutorial https://m6anet.readthedocs.io/en/latest/quickstart.html

However, when I have a look at the output file, instead of indicating a 0 or a 1 as shown in the examples, mine turns out to be a blank column.

For your reference I have attached a snippet of my output file:

transcript_id, transcript_position, n_reads, probability_modified, kmer 2L, 74989, 31, ,TGACT 2L, 75008, 32, ,AAACT 2L, 75100, 33, ,TAACA 2L, 75106, 31, ,TGACT 2L, 75151, 35, ,AAACT

If you can provide me some guidance with this that will be great. Thank you!

[Stakaitis]

Your _nreads column is also empty which is probably the initial problem. Could you show the head of m6anet-dataprep outputs:

• data.log
• data.readcount
• eventalign.index
• data.index

[BioinfoHub-PeiQinNg]

Hi there, Thank you for getting back to me.

These are what the few output head of m6anet-dataprep looks like

• head data.log 2L: Data preparation ... Done. 2R: Data preparation ... Done. 3L: Data preparation ... Done. 3R: Data preparation ... Done. 4: Data preparation ... Done. X: Data preparation ... Done. Y: Data preparation ... Done. dmel_mitochondrion_genome: Data preparation ... Done. Unmapped_Scaffold_8: Data preparation ... Done. 3Cen_mapped_Scaffold_31: Data preparation ... Done.

• head data.readcount transcript_id,transcript_position,n_reads 2L,49675,1 2L,51533,1 2L,51692,1 2L,51748,1 2L,51783,1 2L,51792,1 2L,51883,2 2L,51927,1 2L,52054,2

• eventalign.index transcript_id,read_index,pos_start,pos_end 2L,0,154,56988 2L,1,56988,117016 2L,2,117016,229586 2L,3,229586,470294 2L,5,470294,934066 2L,6,934066,1334426 2L,7,1334426,1873408 2L,8,1873408,2155664 2L,9,2155664,2304450

• head data.index transcript_id,transcript_position,start,end 2L,49675,0,69 2L,51533,69,138 2L,51692,138,207 2L,51748,207,276 2L,51783,276,345 2L,51792,345,414 2L,51883,414,521 2L,51927,521,590 2L,52054,590,697

Thank you again for assisting me with this.

[BioinfoHub-PeiQinNg]

I have also provided the command lines that I have used for nanopore eventalign and m6a-dataprep for your reference:

nanopolish eventalign \ --reads {input.fasta} \ --bam {input.bam} \ --genome {input.ref} \ --scale-events > {SAMPLE}-ref.eventalign.txt

m6anet-dataprep --eventalign {input.txt} \ --out_dir {output} \

[chrishendra93]

Thank you @Stakaitis for helping to respond. @BioinfoHub-PeiQinNg your nanopolish eventalign is missing --signal-index argument which tells m6Anet the segment length information

[BioinfoHub-PeiQinNg]

I see. I will give it another go. Thank you so much!

[chrishendra93]

hi @BioinfoHub-PeiQinNg, how is it working with you? Are you able to get the results now?

[BioinfoHub-PeiQinNg]

Hi @chrishendra93 , sorry for the late reply. I manage to get it running now! Thank you so much for your help! Cheers ^^