add complete conda installation and scripts for ecDNA processing

Author: Madalina Giurgiu

genomics/circle-seq/README.md

@@ -18,12 +18,8 @@ Run the analysis for the PDX primary and relapse:
 ```
 # setup variables
 REF=../genome/hs37d5_mm10.fa
-PRIMARY_SAMPLE=PDXprimary
-RELAPSE_SAMPLE=PDXrelapse
-PRIMARY_1_FASTQ=ecDNA/lane1PrimaryPDX_1_sequence.fq.gz
-PRIMARY_2_FASTQ=ecDNA/lane1PrimaryPDX_2_sequence.fq.gz
-RELAPSE_1_FASTQ=ecDNA/lane1RelapsePDX_1_sequence.fq.gz
-RELAPSE_2_FASTQ=ecDNA/lane1RelapsePDX_2_sequence.fq.gz
+PRIMARY_SAMPLE=PrimaryPDX
+RELAPSE_SAMPLE=RelapsePDX
 
 # run analysis
 bash run.sh

genomics/circle-seq/bwa_mem_align.sh

@@ -4,22 +4,41 @@ date
 hostname
 
 sample=$1
-read1=$2
-read2=$3
-genome=$4
+genome=$2
 
 ##get quality of fastqs
-mkdir -p ${sample}/qc/
-fastqc -o ${sample}/qc/ ${read1} ${read2}
+mkdir -p ecDNA/${sample}/qc/
+fastqc -o ecDNA/${sample}/qc/ ecDNA/${sample}/lane1${sample}_1_sequence.fq.gz ecDNA/${sample}/lane1${sample}_2_sequence.fq.gz
+
+##trim reads
+trim_galore -q 20 --phred33 --illumina --paired -j 4 ecDNA/${sample}/lane1${sample}_1_sequence.fq.gz ecDNA/${sample}/lane1${sample}_2_sequence.fq.gz
+mv lane1${sample}_1_sequence.fq.gz_trimming_report.txt ecDNA/${sample}/
+mv lane1${sample}_1_sequence_val_1.fq.gz ecDNA/${sample}/
+mv lane1${sample}_2_sequence.fq.gz_trimming_report.txt ecDNA/${sample}/
+mv lane1${sample}_2_sequence_val_2.fq.gz ecDNA/${sample}/
 
 ##align with bwa mem
-bwa mem -q -t 4 ${genome} ${read1} ${read2} | samtools sort -@4 -o sorted_${sample}.bam -
+bwa mem -q -t 4 ${genome} ecDNA/${sample}/lane1${sample}_1_sequence_val_1.fq.gz ecDNA/${sample}/lane1${sample}_2_sequence_val_2.fq.gz | samtools sort -@4 -o ecDNA/${sample}/sorted_${sample}.bam -
+
+##index bam file
+samtools index ecDNA/${sample}/sorted_${sample}.bam
+
+##mark duplicates
+picard MarkDuplicates \
+	I=ecDNA/${sample}/sorted_${sample}.bam \
+	O=ecDNA/${sample}/sorted_${sample}.dedup.bam \
+	M=ecDNA/${sample}/sorted_${sample}.dedup.metrix
 
 ##index bam file
-samtools index sorted_${sample}.bam
+samtools index ecDNA/${sample}/sorted_${sample}.dedup.bam
 
 ##create coverage overview in bigwig format
-bamCoverage --bam sorted_${sample}.bam -o ${sample}.bw
+bamCoverage --bam ecDNA/${sample}/sorted_${sample}.dedup.bam -o ecDNA/${sample}/sorted_${sample}.dedup.bw
+
+## remove files
+rm ecDNA/${sample}/sorted_${sample}.bam*
+
+chmod +x ecDNA/${sample}/*
 
 echo "=========================================================="
 echo "Finished on : $(date)"

genomics/circle-seq/circleseq.yaml

@@ -10,4 +10,7 @@ dependencies:
   - openjdk==11.0.1
   - picard==2.25.0
   - python==2.7.15
-  - bwamem==0.7.17
+  - bwa==0.7.17
+  - fastqc==0.11.9
+  - pysam==0.14.1
+  - trim-galore==0.6.7

genomics/circle-seq/run.sh

@@ -1,18 +1,28 @@
 #!/bin/bash
+#SBATCH --job-name=circleseq
+#SBATCH --ntasks=16
+#SBATCH --nodes=1
+#SBATCH --time=1-00:00:00
+#SBATCH --mem=30G
 
 date
 hostname
 
-conda activate circleseq
+#conda activate circleseq
+
+PRIMARY_SAMPLE=PrimaryPDX
+RELAPSE_SAMPLE=RelapsePDX
 
 # mapping
-bash bwa_mem_align.sh $PRIMARY_SAMPLE $PRIMARY_1_FASTQ $PRIMARY_2_FASTQ $REF
-bash bwa_mem_align.sh $PRIMARY_SAMPLE $RELAPSE_1_FASTQ $RELAPSE_2_FASTQ $REF
+#bash bwa_mem_align.sh $PRIMARY_SAMPLE $REF
+#bash bwa_mem_align.sh $RELAPSE_SAMPLE $REF
 
 # circle-enrich-finder
-git clone git clone https://github.com/henssen-lab/circle-enrich-filter.git
+#git clone https://github.com/henssen-lab/circle-enrich-filter.git
+# checkout version 1.0.0
 cd circle-enrich-filter
-bash run_CircleEnrichFilter.sh $PRIMARY_SAMPLE/sorted_PDXprimary.dedup.bam $PRIMARY_SAMPLE/circleEnrich0 $REF
-bash run_CircleEnrichFilter.sh $RELAPSE_SAMPLE/sorted_PDXprimary.dedup.bam $RELAPSE_SAMPLE/circleEnrich0 $REF
+git checkout f0fc687c974c9e4308e423f9cbe883344c6b359a
+bash run_CircleEnrichFilter.sh -i ${PWD}/../ecDNA/$PRIMARY_SAMPLE/sorted_${PRIMARY_SAMPLE}.dedup.bam -o ${PWD}/../ecDNA/$PRIMARY_SAMPLE/circleEnrich0 -s 0
+bash run_CircleEnrichFilter.sh -i ${PWD}/../ecDNA/$RELAPSE_SAMPLE/sorted_${RELAPSE_SAMPLE}.dedup.bam -o ${PWD}/../ecDNA/$RELAPSE_SAMPLE/circleEnrich0 -s 0
 
-conda deactivate
+#conda deactivate