Pipeline for discovering splice-site creating mutations (SCMs) from RNA-Seq. It works on LSF job scheduler and can run multiple jobs in parallel. For other job submission system, please modify the script accordingly.
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Cao S, Zhou DC, Oh C, Jayasinghe RG, Zhao Y, Yoon CJ, Wyczalkowski MA, Bailey MH, Tsou T, Gao Q, Malone A, Reynolds S, Shmulevich I, Wendl MC, Chen F, Ding L. Discovery of Driver Non-Coding Splice-Site-Creating Mutations in Cancer. Nat Commun 2020 Nov 4;11(1):5573
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Jayasinghe RG*, Cao S*, Gao Q, Wendl MC, Vo NS, Reynolds SM, Zhao Y, Climente-González H, Chai S, Wang F, Varghese R, Huang M, Liang W, Wyczalkowski MA, Sengupta S, Li Z, Payne SH, Fenyö D, Miner JH, Walter MJ, The Cancer Genome Atlas Research Network, Vincent B, Eyras E, Chen K, Shmulevich I, Chen F, Ding L, Systematic Analysis of Splice-Site-Creating Mutations in Cancer, Cell Rep 2018 Apr 3;23(1):270-281
Basic Linux:
sudo apt-get install -y samtools.
With Conda environment:
conda install -c bioconda samtools
perl misplice.pl --rdir run_folder --bed bed_file --ref ref_file --q q_name --maf f_maf --bamlist f_bam --n n_file --step step_number
<run_folder> = full path of the folder holding mutation annotation file (maf) file for all mutations (misplice.input.maf) and file for the sample list (currently named as Samples).
<bed_file> bed file for HG38 refernece: please follow ./resource/GRCh38/work_log_gtf to generate it or download from https://drive.google.com/file/d/1M2yLLjxBwOjUOu-uIxwtUrD_DHTTpioJ/view?usp=sharing
bed file for HG19 reference can be downloaded from https://drive.google.com/file/d/14vHI3G45c-xBzpgzs_I43orrKbNJyIVi/view?usp=sharing
<ref_file> HG38 or HG19 refernece file
<q_name> which bsub quenue for submitting job
<f_maf> Somatic mutation file (in maf format), standard output from vcf2maf script
Example can be found from misplice.example.maf
<f_bam> file for input bam list
Example can be found from misplice.rnabam.tsv. Make sure the sample name in bam list file is matching the tumor name in misplice.example.maf file.
<n_file> number of split files for maf file: Default 100.
Example can be found from misplice.rnabam.tsv, which contains four columns (sample_name, cancer_type, rna_bam_path, and chr_status).
chr_status indicates whether the chr in the bam contains chr or not. If it has chr, type chr1 in the four column. Otherwise, type 1.
** Make sure the sample_name is consistent with tumor name in the maf file: tumor name in the maf file with an additional suffix _T is fine if variants are called by somaticwrapper. It will be automatically removed by the pipeline. **
<step_number> run this pipeline step by step
[1] Split mutation annotation file (MAF) into multiple files for processing
[2] Run discovery step of mutation induced splice creating events
[3] Generate control MAF
[4] Calculate number of supporting reads for control samples
[5] Combine the read count for case and control samples
[6] Calculate the splice score (MaxEntScan)
[7] Split novel sites for bam readcounts
[8] Do bam readcounts
[9] Run rc, hla and expression filtering
Song Cao, Reyka Jayasinghe, Qingsong Gao, Mike Wendl, Li Ding
Pipeline leading contact: scao@wustl.edu