MET exon 14 skipping

[sotaro kanematsu]

I am considering using rMATS to detect tumor-specific splicing events. As a first step, I will test whether rMATS can detect exon skipping by using the BAM file of a sample in which MET exon 14 skipping was observed in the tumor region.Unfortunately, MET exon 14 skipping is still not detected even with the --statoff option. I sequenced a strand-specific RNA-seq library and used the aligned BAM file from STAR as the input. What could be the cause of this issue? I am using rMATS version 4.3.0.

(rmats) [root@localhost splicing]# time sh ../script/rmats_statoff.sh
gtf: 31.105498552322388
There are 57820 distinct gene ID in the gtf file
There are 196520 distinct transcript ID in the gtf file
There are 35613 one-transcript genes in the gtf file
There are 1196293 exons in the gtf file
There are 24864 one-exon transcripts in the gtf file
There are 21893 one-transcript genes with only one exon in the transcript
Average number of transcripts per gene is 3.398824
Average number of exons per transcript is 6.087386
Average number of exons per transcript excluding one-exon tx is 6.824282
Average number of gene per geneGroup is 7.482150
statistic: 0.03349876403808594

read outcome totals across all BAMs
USED: 0
NOT_PAIRED: 1492375
NOT_NH_1: 18348078
NOT_EXPECTED_CIGAR: 2066313
NOT_EXPECTED_READ_LENGTH: 158519928
NOT_EXPECTED_STRAND: 0
EXON_NOT_MATCHED_TO_ANNOTATION: 0
JUNCTION_NOT_MATCHED_TO_ANNOTATION: 0
CLIPPED: 14602821
total: 195029515
outcomes by BAM written to: /run/media/skanematsu/kanematsu/RNASeq/splicing/rmats_result_statoff/tmp/2024-11-06-23_57_14_417636_read_outcomes_by_bam.txt

novel: 486.0649039745331
The splicing graph and candidate read have been saved into /run/media/skanematsu/kanematsu/RNASeq/splicing/rmats_result_statoff/tmp/2024-11-06-23_57_14_417636_*.rmats
save: 0.00033926963806152344
loadsg: 0.0006213188171386719

==========
Done processing each gene from dictionary to compile AS events
Found 38132 exon skipping events
Found 2040 exon MX events
Found 12653 alt SS events
There are 7727 alt 3 SS events and 4926 alt 5 SS events.
Found 5558 RI events
==========

ase: 1.6592433452606201
count: 0.2392866611480713
Processing count files.
Done processing count files.

real 8m42.668s
user 16m22.078s
sys 0m5.606s

file:///home/skanematsu/METexon14.png

[sotaro kanematsu]

Here is the script I created.

#!/bin/bash

RMATS="/home/skanematsu/anaconda3/envs/rmats/bin/rmats.py"


# 参照アノテーションのGTFファイル
GTF="/run/media/skanematsu/kanematsu/RNASeq/arriba/GENCODE19.gtf"

# 出力ディレクトリ
OUTPUT_DIR="/run/media/skanematsu/kanematsu/RNASeq/splicing/rmats_result_statoff"

mkdir -p ${OUTPUT_DIR}

# スレッド数の指定
THREADS=8

# rmatsの実行
$RMATS          --b1 normal_bams.txt \
                --b2 tumor_bams.txt \
                --gtf $GTF \
                --od $OUTPUT_DIR \
                --tmp $OUTPUT_DIR/tmp \
                -t paired \
                --readLength 145 \
                --statoff  \
                --nthread $THREADS \
                --libType fr-firststrand

2024年11月7日木曜日 8:46:50 UTC+9 sotaro kanematsu:

[sotaro kanematsu]

No problem at all! It turns out it was a mistake with the readLength setting.

Is it possible to specify sample names in the header of the rMATS results? Currently, they appear as SAMPLE_1, SAMPLE_2, etc.

2024年11月7日木曜日 8:49:10 UTC+9 sotaro kanematsu: