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Whole-Genome Deoxyribonucleic Acid Extraction from Mycobacterium Species via the Cetyltrimethylammonium Bromide Technique
Published on: December 12, 2025
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Complete nontuberculous mycobacteria whole genomes using an optimized DNA extraction protocol for long-read
Jennifer M Bouso1, Paul J Planet2,3,4
1Division of Pulmonary Medicine, Children's Hospital of Philadelphia, Philadelphia, PA, USA.
BMC Genomics
|November 1, 2019
Summary
A new DNA extraction protocol efficiently yields high-quality genomic DNA from nontuberculous mycobacteria (NTM) for long-read whole genome sequencing (WGS). This method facilitates complete NTM genome assembly and aids in understanding NTM evolution and pathogenicity.
Area of Science:
- Microbiology
- Genomics
- Molecular Biology
Background:
- Nontuberculous mycobacteria (NTM) cause significant pulmonary and systemic diseases.
- Genomic research on NTM transmission, evolution, and pathogenicity is increasing.
- Extracting pure genomic DNA from NTM is challenging due to slow growth and tough cell walls.
Purpose of the Study:
- To develop an optimized DNA extraction protocol for long-read whole genome sequencing (WGS) of NTM.
- To obtain large quantities of highly pure genomic DNA from NTM without additional clean-up steps.
Main Methods:
- Compared the optimized DNA extraction method with 6 other protocols.
- Tested the method on 38 clinical isolates from M. avium and M. abscessus complexes.
- Utilized Oxford Nanopore Technologies and MinION for long-read sequencing.
Main Results:
- The optimized protocol yielded optimal quality and quantity DNA for long-read WGS.
- Successfully completed circularized M. avium subspecies hominissuis genomes.
- Identified a novel plasmid within M. avium subspecies hominissuis.
Conclusions:
- The optimized DNA extraction protocol and assembly pipeline are efficient for complete mycobacterial genome closure.
- This method is a valuable tool for long-read sequencing and assembly of mycobacterial genomes.
- Long-read sequencing approaches will advance the understanding of NTM evolution and pathogenicity.
