Mycobacterium tuberculosis is a human pathogen, requiring
specialist containment facilities (BSL-3, CL-3, PC3); all handling
should be performed under the local regulations for your region.
2.1 Sputum
Decontamination
and DNA Extraction &
Quantification
1. N-acetyl-L-cysteine/NAOH (1% NaOH final concentration).
2. QIAamp Mini Kit (Qiagen).
3. DNA LoBind 1.5 mL tubes (Eppendorf).
4. PBS (sterile).
5. Vortex mixer.
6. 1.5 mL/2 mL tube heat block.
7. 2 mL screw cap tubes.
8. 0.1 mm glass beads (Beckton Dickenson).
9. Microfuge.
10. 96–100% ethanol, molecular biology grade.
11. Nuclease-free water.
12. Qubit dsDNA HS Assay Kit, and Qubit 3.0 Fluorometer (Life
Technologies).
13. Qubit Assay Tubes (Life Technologies).
14. 50 mL sterile Falcon tubes (Corning).
Fig. 1 Schematic of the library preparation steps
462
Amanda Claire Brown
specialist containment facilities (BSL-3, CL-3, PC3); all handling
should be performed under the local regulations for your region.
2.1 Sputum
Decontamination
and DNA Extraction &
Quantification
1. N-acetyl-L-cysteine/NAOH (1% NaOH final concentration).
2. QIAamp Mini Kit (Qiagen).
3. DNA LoBind 1.5 mL tubes (Eppendorf).
4. PBS (sterile).
5. Vortex mixer.
6. 1.5 mL/2 mL tube heat block.
7. 2 mL screw cap tubes.
8. 0.1 mm glass beads (Beckton Dickenson).
9. Microfuge.
10. 96–100% ethanol, molecular biology grade.
11. Nuclease-free water.
12. Qubit dsDNA HS Assay Kit, and Qubit 3.0 Fluorometer (Life
Technologies).
13. Qubit Assay Tubes (Life Technologies).
14. 50 mL sterile Falcon tubes (Corning).
Fig. 1 Schematic of the library preparation steps
462
Amanda Claire Brown