Course: BIOL 4150 / BIOL 6150
Overview
This project focuses on aligning post-QC sequencing reads (FASTQ files) to the human reference genome (GRCh38). Key objectives include preparing the reference genome, aligning reads using BWA-MEM2, and processing the results using SAMtools. The analysis provides practical experience in bioinformatics workflows for next-generation sequencing (NGS) data.
Repository Link: https://github.com/ilahamusayeva93/Project-2-Read-Mapping
Project Details
Estimated Time: ~1.5 hours Tools Used: BWA-MEM2: For aligning sequencing reads to the reference genome. SAMtools: For sorting, conversion, and evaluating alignment files. Environment Configuration Processor: Intel(R) Xeon(R) Gold 6226 CPU @ 2.70GHz Cores: 24 (12 cores per socket, 2 sockets) Memory: 754 GB Storage: 50 TB (network-mounted) and 1.4 TB (local) Repository Contents
Project2-ReadMapping.ipynb: Starter notebook with detailed steps for read mapping and post-alignment analysis. Input Files: Post-QC FASTQ Files: SRR081224_1_trimmed.fastq.gz SRR081224_2_trimmed.fastq.gz Reference Genome: hg38.fa (GRCh38 human reference genome) Scripts and Outputs: Example commands for reference genome indexing, read alignment, and SAMtools-based file processing. Intermediate and final output files, including: Indexed reference genome files (.amb, .ann, .bwt, .fai, etc.) SAM file: SRR081224.sam Sorted SAM file: SRR081224_sorted.sam BAM file: SRR081224.bam Objectives
Prepare Reference Genome Download and index the GRCh38 genome for alignment. Understand the meaning of uppercase/lowercase nucleotides and gaps (N) in FASTA files. Align Sequencing Reads Map paired-end reads to the indexed reference genome using BWA-MEM2. Optimize computational resource usage with SLURM-based batch jobs. Evaluate Alignment Quality Process SAM files: sorting, conversion to BAM, and statistical evaluation. Analyze individual reads (e.g., CIGAR string, chromosome alignment) for detailed insights. How to Use
Clone the repository: git clone https://github.com/ilahamusayeva93/Project-2-Read-Mapping cd Project-2-Read-Mapping Ensure all dependencies are installed: BWA-MEM2 SAMtools Follow the steps in the Project2-ReadMapping.ipynb notebook: Prepare the reference genome. Perform read alignment. Process and evaluate the outputs. Expected Outputs
Intermediate Files: Indexed Reference Genome: (hg38.fa, .amb, .ann, .bwt, .pac, .fai) SAM File: SRR081224.sam Sorted SAM File: SRR081224_sorted.sam BAM File: SRR081224.bam Key Insights: Total raw and mapped reads. Detailed analysis of alignment quality and specific read alignments. Acknowledgments
This project is part of Georgia Tech's BIOL 4150 / BIOL 6150 coursework. Computational resources were provided by the PACE-ICE cluster.