Small snippet that generates a genome index and aligns the RNAseq reads.

rnaseq/step3_align/snippet.sh

@@ -0,0 +1,53 @@
+#!/bin/bash
+#
+# Align reads against reference genome.
+
+STORAGE="/groups/umcg-griac/tmp04/rawdata/$(whoami)/step3"
+# Store genome index in this location:.
+GENOME_INDEX="${STORAGE}/genome_index"
+mkdir -p "${GENOME_INDEX}"
+# Store the generated `Aligned.sortedByCoord.out.bam` in this dir.
+ALIGNMENT_OUTPUT="${STORAGE}/alignment"
+mkdir -p "${ALIGNMENT_OUTPUT}"
+
+# 1) Generate genome index.
+#
+# N.B.:
+# - We're assuming a read size of 100 bp (--sjdbOverhang 100). Refer back to the
+#   previous quality control steps if you are unsure about the size. In case of
+#   reads of varying length, the ideal value is max(ReadLength)-1.
+# - We're using gzip compressed reference data (--readFilesCommand zcat), i.e.,
+#   .gtf.gz and fa.gz. If not, you can remove the `zcat` flag.
+
+# Storage location reference data (in this case on calculon).
+REFERENCE_DATA="/groups/umcg-griac/prm02/rawdata/reference/genome"
+GTF_FILE="${REFERENCE_DATA}/Homo_sapiens.GRCh38.100.gtf.gz"
+FASTA_FILE="${REFERENCE_DATA}/Homo_sapiens.GRCh38.dna.primary_assembly.fa.gz"
+
+STAR \
+    --runThreadN 8 \
+    --runMode genomeGenerate \
+    --readFilesCommand zcat \
+    --sjdbOverhang 100 \
+    --genomeFastaFiles ${FASTA_FILE} \
+    --sjdbGTFfile ${GTF_FILE} \
+    --genomeDir ${GENOME_INDEX}
+
+
+# 2) Do the actual alignment.
+#
+# N.B.:
+# - We are assuming paired-end, gzip compressed (--readFilesCommand zcat)  FastQ
+#   files.
+
+# THe compressed paired-end FastQ's that we are aligning.
+R1="sample1_R1.fastq.gz"
+R2="sample1_R2.fastq.gz"
+
+STAR \
+    --runThreadN 8 \
+    --readFilesCommand zcat \
+    --readFilesIn "${R1}" "${R2}" \
+    --outSAMtype BAM SortedByCoordinate \
+    --genomeDir ${GENOME_INDEX} \
+    --outFileNamePrefix "${ALIGNMENT_OUTPUT}"

rnaseq/step5_count/snippet_counts

@@ -1,18 +0,0 @@
-## Generate Raw Counts using FeatureCounts/HTSeqCounts 
-
-## Load package Subread (packge within which FeatureCounts is present)
-
-+ ml Subread/1.6.4-foss-2015b FeatureCounts
-
-## Features used in FeatureCounts : 
-## -s : reversely stranded 
-## -T : number of threads used 
-## -a : reference genome 
-## -o : output file 
-
-+ featureCounts \
-+ --a "path to reference genome" \
-+ --T 6 \
-+ --o "path to output file" \
-+ --s 2
-