#!/bin/bash
#
# Align reads against reference genome.

REFERENCE_DATA="/groups/umcg-griac/prm03/rawdata/reference/genome"

PROJECT_DIRECTORY="/groups/umcg-griac/tmp01/rawdata/$(whoami)/rnaseq"
# Store the generated `sample1_Aligned.sortedByCoord.out.bam` in this dir.
ALIGNMENT_OUTPUT="${PROJECT_DIRECTORY}/step3/alignment"
mkdir -p "${ALIGNMENT_OUTPUT}"

# 1) Generate genome index (optional).
#
# Depending on your read size, reference genome and annotation, you may need to
# generate a new genome index. In most cases, this is not necessary and you can
# directly use the pre-build genome index from the cluster:
#
# GENOME_INDEX="${REFERENCE_DATA}/index_GRCh38_gtf100_overhang100"
#
# and ignore the first STAR command below.
#
# N.B.:
# - We're assuming a read size of 100 bp (--sjdbOverhang 100). An alternative
#   cut-off is 150, for low-input methods. In general, refer back to the
#   previous quality control steps if you are unsure about the size. In case of
#   reads of varying length, the ideal value is max(ReadLength)-1.
# - If you're using gzip compressed reference data, i.e., .gtf.gz and fa.gz,
#   pass the `--readFilesCommand zcat` flag.

# Store created genome index in this location:.
GENOME_INDEX="${PROJECT_DIRECTORY}/step3/genome_index"
mkdir -p "${GENOME_INDEX}"

# Storage location reference data on Gearshift.
GTF_FILE="${REFERENCE_DATA}/Homo_sapiens.GRCh38.100.gtf"
FASTA_FILE="${REFERENCE_DATA}/Homo_sapiens.GRCh38.dna.primary_assembly.fa"

STAR \
    --runThreadN 8 \
    --runMode genomeGenerate \
    --sjdbOverhang 100 \
    --genomeFastaFiles ${FASTA_FILE} \
    --sjdbGTFfile ${GTF_FILE} \
    --genomeDir ${GENOME_INDEX}


# 2) Do the actual alignment.
#
# N.B.:
# - We are assuming paired-end, gzip compressed (--readFilesCommand zcat)  FastQ
#   files.

# The compressed, paired-end, FastQ's after trimming (step 2).
R1="${PROJECT_DIRECTORY}/step2/sample1_R1_paired.fastq.gz"
R2="${PROJECT_DIRECTORY}/step2/sample1_R2_paired.fastq.gz"

STAR \
    --runThreadN 8 \
    --readFilesCommand zcat \
    --readFilesIn "${R1}" "${R2}" \
    --outSAMtype BAM SortedByCoordinate \
    --genomeDir ${GENOME_INDEX} \
    --outFileNamePrefix "${ALIGNMENT_OUTPUT}/sample1_"