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nloyfer / UXM_deconv

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Nanopore Data #15

Open AzlanNI opened 7 months ago

AzlanNI commented 7 months ago

Hello everyone!

My Name is Azlan and i am currently analyzing our ONT Data. I wanted to use uxm and wgbs_tools to further investigate the methylation status in different samples and just wanted to ask if i can use these tools for ONT. I read a lot about wgbs Data and just am curious if anyone has done deconvolution and methylation analysis for Nanopore Data with these Tools.

Kind regards, Azlan

AzlanNI commented 7 months ago

So i know tried out a lot and got to a Problem. I am using Guppy for basecalling and generating .bam files. After that i used wgbstool to generate .pat and .beta files from the .bam file. Now i am trying to use the .pat File i got from wgbstools to deconvolve with uxm. So my command ist uxm deconv Barcode02_sorted.pat.gz -o Output.csv And it is giving me: Invalid input argument Length of values (2) does not match length of index (36)

I tried the following: I got the markers from tail -n +2 Atlas.U250.l4.hg19.full.tsv | cut -f1-5 > markers_Atlas.U250.l4.hg19.full.bed And used wgbstool view -L to filter my pat file wgbstools view -L /Users/azlannisar/UXM_deconv/supplemental/markers_Atlas.U250.l4.hg19.full.bed Barcode02_sorted.pat.gz --min_len 4 --strip --strict > Barcode02_Nano_sorted.stripped.pat

After that my File is empty so i guess i dont have any Regions which are overlapping ? So how should i go on can i somehow adjust the marker file or is it not possible for Nanopore Data.

I am thankful for any help! kind regards, Azlan

AzlanNI commented 7 months ago

after using wgbstools homog -b blocks_Barcode02.bed.gz -f Barcode02_sorted.pat.gz i get WARNING: all zeros! But i generated the files from the same .bam file I looked more into it and generated the blocks file with this command: wgbstools segment --genome GENOME_NAME -F Barcode02_sorted_ascii.beta -o blocks_barcode02_new.bed The GENOME_NAME file is the same reference i generated the bam file with. And if i look into the first entries of the pat, beta and blocks file it looks like this Pat : (base) azlannisar@Air-von-Azlan Desktop % head -n 10 Barcode02_sorted.pat 1 236 TCCT.CCCCCCCCCCCCC.C 4 1 1166 CTTTC.T 4 1 1187 TTTTCCCCCCC 4 1 1194 CCCCT.CCCCCTCCCCCCCCCCCCCCCC 4 1 2046 TTCCCCTCT 4 1 2048 C.CCCC....CCC..C.T.TCCCCTCCCC...C.CC..CTCC 4 1 2133 CTT.C.TCCCCT.CCC.CCC 4 1 2153 C..C.TCC..CCCCCTT.CCCCCCC..C.CT.CCCC.T.CTC 4 1 2192 C 4 1 4067 TCCCC.CCCCCCCCCCCCCCCCTT..TTTTTTCCCCCCCCCCCCCCCCC.CCCCCCCT.TTCTCC.CCCCCCCCCCCTCC.C.C.T.CCCCTCCCCCCC 4 beta: (base) azlannisar@Air-von-Azlan Desktop % head -n 10 Barcode02_sorted_beta.bed 1 16821 16823 0 4 1 16868 16870 4 4 1 16931 16933 4 4 1 16943 16945 0 4 1 16973 16975 4 4 1 17377 17379 4 4 1 17405 17407 4 4 1 17451 17453 4 4 1 17477 17479 4 4 1 17482 17484 4 4 Blocks: head -n 10 blocks_barcode02_new.bed 1 10469 10782 1 55 1 10783 12783 55 154 1 12807 14792 154 190 1 14888 16823 190 237 1 16962 17716 240 256 1 17853 19789 256 304 1 19888 21818 304 348 1 21948 23844 348 375 1 23974 25934 375 403 1 25974 27970 403 430 So i guess my mistake is in the segmentation if i am not mistaken. Do i have to toggle something like in bam2pat with. -np for nanopore Data ?