dmr multi failed to load samples

[nickst85]

Hi,

I'm trying to run dmr multi to compare bedMethyl pileup files from three patients and three controls.
the code is as folows:

modkit dmr multi
-s $INPATH/patient1_pileup.bed.gz patient1
-s $INPATH/patient2_pileup.bed.gz patient2
-s $INPATH/patient3_pileup.bed.gz patient3
-s $INPATH/Control1_pileup.bed.gz Control1
-s $INPATH/Control2_pileup.bed.gz Control2
-s $INPATH/Control3_pileup.bed.gz Control3
-i $INPATH/patient1_pileup.bed.gz.tbi patient1
-i $INPATH/patient2_pileup.bed.gz.tbi patient2
-i $INPATH/patient3_pileup.bed.gz.tbi patient3
-i $INPATH/Control1_pileup.bed.gz.tbi Control1
-i $INPATH/Control2_pileup.bed.gz.tbi Control2
-i $INPATH/Control3_pileup.bed.gz.tbi Control3
--regions-bed $REFERENCE/hg38_v44.bed
--ref $REFERENCE/gencode.v44.transcripts.fa
--out-dir $OUTPATH/patient_vs_controls_dmr_multi.bed
--base A
-t 80

when I run it I get the following error file:

failed to load patient1
failed to load patient2
failed to load patient3
failed to load Control1
failed to load Control2
failed to load Control3
loaded 252835 regions
processing 120 regions concurrently
Error! n must be > 2

the pileup files look like:

ENST00000655252.1 119 120 a 2 + 119 120 255,0,0 2 0.00 0 2 0 0 0 0 0
ENST00000655252.1 226 227 a 2 + 226 227 255,0,0 2 0.00 0 2 0 0 1 0 0
ENST00000655252.1 230 231 a 2 + 230 231 255,0,0 2 100.00 2 0 0 0 1 0 0
ENST00000655252.1 268 269 a 2 + 268 269 255,0,0 2 0.00 0 2 0 0 0 1 0
ENST00000655252.1 344 345 a 3 + 344 345 255,0,0 3 100.00 3 0 0 0 0 0 0
ENST00000655252.1 492 493 a 3 + 492 493 255,0,0 3 0.00 0 3 0 0 0 0 0
ENST00000655252.1 787 788 a 3 + 787 788 255,0,0 3 0.00 0 3 0 0 0 0 0
ENST00000655252.1 794 795 a 3 + 794 795 255,0,0 3 0.00 0 3 0 0 0 0 0
ENST00000655252.1 1003 1004 a 4 + 1003 1004 255,0,0 4 0.00 0 4 0 0 0 0 0
ENST00000655252.1 1012 1013 a 4 + 1012 1013 255,0,0 4 0.00 0 4 0 0 0 0 0

What's happening?

Thank you in advance,

Nick

[ArtRand]

Hello @nickst85,

Could you add --log-filepath debug.log to your command and paste the content of the file here? This error originates from modkit failing to find either the bedmethyl or index files.

[nickst85]

Hi Arthur, Thank you for your prompt response. Please see attached the debug.log file. Let me know what I should modify. Thank you in advance, Nick From: Arthur Rand ***@***.***> Date: Thursday, June 13, 2024 at 7:48 PM To: nanoporetech/modkit ***@***.***> Cc: Settas, Nikolaos ***@***.***>, Mention ***@***.***> Subject: [EXT] Re: [nanoporetech/modkit] dmr multi failed to load samples (Issue #207) ATTENTION: External Email! Do not click attachments/links unless sender is known.

…

________________________________ Hello @nickst85 [github. com], Could you add --log-filepath debug. log to your command and tell me what it says? This error originates from modkit failing to find either the bedmethyl or index files. — Reply to this email directly, view Hello @nickst85 [github.com]<https://urldefense.com/v3/__https:/github.com/nickst85__;!!FMDslA!YBA-qzjd-XgEcjHf6wZC-eeiJQJ-wZ10AZi7Z4o6VBeI4J62C_a_tf6rXX-F70dZj0DCoNniLxiv1e0mKAjGY4pbEG6F4Yw$>, Could you add --log-filepath debug.log to your command and tell me what it says? This error originates from modkit failing to find either the bedmethyl or index files. — Reply to this email directly, view it on GitHub [github.com]<https://urldefense.com/v3/__https:/github.com/nanoporetech/modkit/issues/207*issuecomment-2166968470__;Iw!!FMDslA!YBA-qzjd-XgEcjHf6wZC-eeiJQJ-wZ10AZi7Z4o6VBeI4J62C_a_tf6rXX-F70dZj0DCoNniLxiv1e0mKAjGY4pbmT9Uapc$>, or unsubscribe [github.com]<https://urldefense.com/v3/__https:/github.com/notifications/unsubscribe-auth/A5QGITX6W2I4ZVURRNQBJU3ZHIVUZAVCNFSM6AAAAABJI5TI2OVHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMZDCNRWHE3DQNBXGA__;!!FMDslA!YBA-qzjd-XgEcjHf6wZC-eeiJQJ-wZ10AZi7Z4o6VBeI4J62C_a_tf6rXX-F70dZj0DCoNniLxiv1e0mKAjGY4pbmlc1Hio$>. You are receiving this because you were mentioned.Message ID: ***@***.***> Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message.

[ArtRand]

Hello @nickst85,

It doesn't look like attachments to emails make it through to github issue comments. Could you attach it through github.com or email it to me art.rand[at]nanoporetech.com

[ArtRand]

Hello @nickst85

Thanks for sending the debug log over. As you can tell, there isn't anything that indicates exactly what the problem is in the logs. I've attached a build to this thread that has a little more verbose logging that might help figure out the source of the problem.

Could you run this build and send via email the log or paste it here? It might be worth double checking that all of the paths are valid. Lastly the --out-dir option in your command, --out-dir /home/nsettas/m6A_modkit_dmr/multi/Patients_vs_Controls_dmr_multi.bed, should indicate a directory to write the outputs to, not a single file. The way you have it will still work on most systems, but probably isn't what you intended.
modkit_dev7407fe8_centos7_x86_64.tar.gz

[ArtRand]

Hello @nickst85, any luck?