FASTQ Quality Control and Trimming Guide

Introduction

The raw FASTQ reads sometimes contain adapter sequences and low-quality bases at the ends. These need to be removed before downstream analysis.

Step 1: Install and Use FastQC

Install FastQC:

conda create -n fastqc -c bioconda fastqc

Activate the environment:

conda activate fastqc

Run FastQC on all FASTQ files:

fastqc *.fq.gz

Organize output files:

mkdir zip_files html_files
mv *.zip zip_files/
mv *.html html_files/

To visualize the HTML files:

exit
scp -r name@IP_address:/path/to/html_files ~/

Deactivate FastQC environment:

conda deactivate fastqc

Step 2: Trim Reads with Trim Galore

Install Trim Galore:

conda create -n trim-galore -c bioconda trim-galore

Activate the environment:

conda activate trim-galore

Trim paired-end reads with Illumina adapters:

trim_galore --paired --illumina BEN_CI16_sub_1.fq.gz BEN_CI16_sub_2.fq.gz

View trimming report:

less BEN_CI16_sub_2.fq.gz_trimming_report.txt

Bulk Trimming Options

Option 1: List file names manually

for file in BEN_CI18_sub_1.fq.gz BEN_NW13_sub_1.fq.gz BEN_SI9_sub_1.fq.gz BEN_NW10_sub_1.fq.gz BEN_SI18_sub_1.fq.gz LGS1_sub_1.fq.gz BEN_NW12_sub_1.fq.gz BEN_SI19_sub_1.fq.gz; do
    paired_file=${file/_1.fq.gz/_2.fq.gz}
    trim_galore --paired "$file" "$paired_file"
done

Option 2: Use wildcard to automatically detect file pairs

for file in *_1.fq.gz; do
    trim_galore --paired "$file" "${file/_1.fq.gz/_2.fq.gz}"
done

Output Files:

• *.fq.gz_trimming_report.txt - Summary report.
• *_val_1.fq.gz - Trimmed read 1.
• *_val_2.fq.gz - Trimmed read 2.

Deactivate the Trim Galore environment:

conda deactivate trim-galore