preprocessing_commands.sh

##### Please modify to assign right path to the files accordingly #####

### 1. Processing Long-read ###
## Prepare data
$fa=genome_fasta_file_hg38
$long_fq=long_read_fastq
$te_gtf=TE_individual_gtf

# Minimap2
minimap2 -ax splice $fa $long_fq > processed.sam
samtools view -bS processed.sam > processed.bam

# featureCounts
featureCounts -T 20 -R BAM --verbose -O -L -M -a $te_gtf processed.bam

### 2. Processing Short-read ###
$short_fq1=short_read_fastq_pair1
$short_fq2=short_read_fastq_pair2

## star commands(STAR-2.5.3a Version)
# star index
STAR --genomeChrBinNbits 15 --runMode genomeGenerate --genomeDir STAR_index --genomeFastaFiles $fa --sjdbGTFfile $te_gtf --limitGenomeGenerateRAM 3762779324690
# run star
STAR --runThreadN 20 --genomeDir STAR_index \
   --readFilesIn $short_fq1 $short_fq2 --readFilesCommand zcat --outFileNamePrefix STAR_output \
   --outSAMtype BAM Unsorted --outSAMstrandField intronMotif --outSAMattributes All --outSAMattrIHstart 0 \
   --outFilterMultimapNmax 100 --outFilterScoreMinOverLread 0.4 --outFilterMatchNminOverLread 0.4 --clip3pNbases 0 \
   --winAnchorMultimapNmax 100 --alignEndsType EndToEnd --alignEndsProtrude 100 DiscordantPair --chimSegmentMin 250 \
   --sjdbGTFfile $te_gtf --twopassMode Basic
# sort by read name
samtools collate -o Aligned_sort_name.out.bam --output-fmt BAM Aligned_sort.out.bam

### 3. Generating Simulated Short-read ###
$gtf_combined=Combination_TE+Gene_GTF
# Below link is an actual link to TPM file that our study used
$TPM=https://figshare.com/ndownloader/files/39053627
spankisim_transcripts -g $gtf_combined -f $fa -bp 76 -frag 200 -ends 2 -t $TPM