Development of Rapid Extraction Method of Mycobacterium avium Subspecies paratuberculosis DNA from Bovine Stool Samples
Abstract
:1. Introduction
2. Materials and Methods
2.1. Sample Origin
2.2. Development of MAP SpeedXtract Protocol
2.3. RPA Assay
2.4. Clinical Sensitivity and Specificity
3. Results
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Protocol | Pre-Treatment of the Sample | TT (min) | Exponential Curve | ||||||
---|---|---|---|---|---|---|---|---|---|
10 min; 40 °C | Ultrasonic (20 kHz, 4 min) (BANDELIN electronic, Berlin, Germany) | Bead Beating (1 min) using Soil Grinding SK38 Precellys Tube (Bertin Corp., Rockville, MD, USA) | SpeedXtract Kit (QIAgen Lake Constance, Stockach, Germany) | ||||||
Sodium Dodecyl Sulfate (20 %) 30 µL (Carl Roth, Karlsruhe, Germany) | Proteinase K 60 µL (Carl Roth, Karlsruhe, Germany) | Protease 5 µL (QIAgen, Hilden, Germany) | on Precellys 24 Tissue Homogenizer (6500 rpm) (Bertin Corp., Rockville, MD, USA) | on Vortex (Scientific Industries, Bohemia, NYC, USA) | |||||
1 | + | + | + | + | + | neg | |||
2 | + | + | + | + | 6.0 | + | |||
3 | + | + | + | + | 6.0 | + | |||
4 | + | + | + | neg | |||||
5 | + | + | neg | ||||||
6 | + | + | neg | ||||||
7 | + | + | + | 6.7 | + | ||||
8 | + | + | + | 6.7 | |||||
9 | + | + | 6.3 | + | |||||
10 | + | + | 6.7 | + | |||||
11 | + | neg |
Reference | Kit Used | Kit Producing Company | Purification Method | Time Needed (min) | Sample Amount (mg) | Bead Beating | Heating Step (56 °C–70 °C) | Boiling Step | Proteinase K | Centrifugation | Costs Per reaction (€) |
---|---|---|---|---|---|---|---|---|---|---|---|
Münster et al., 2013 [11] | QIAmp DNA Blood Mini Kit | Qiagen Hilden, Hilden, Germany | silica gel membrane column | 150 | 100 | + | + | + | + | + | 5.90 |
Zang and Zang, 2011 [8] | home-made recipe | silica gel membrane column | 160 | + | + | + | + | ||||
Leite et al., 2013 [16] | MagMax Total Nucleic Acid Isolation Kit | Applied Biosystems, Foster City, CA, USA | magnetic nucleic acid binding beads | 40 | 300 | + | + | 5.52 | |||
PowerSoil DNA Isolation Kit | MO BIO Laboratories Inc., Carlsbad, CA, USA | silica gel membrane column | 40 | 300 | + | + | + | + | 5.00 | ||
QIAamp Stool DNA Mini Kit | Qiagen Hilden, Hilden, Germany | silica gel membrane column | 40 | 1000 | + | + | + | + | + | 5.78 | |
ExtractMaster Fecal DNA Extraction Kit | Epicenter Biotechnologies, Madison, WI, USA | inhibitor removal spin column | 50 | 50 | + | + | + | unknown | |||
ZR Fecal DNA MiniPrep | Zymo Research Corp., Irvine, CA, USA | Spin column | 20 | 150 | + | + | 2.65 | ||||
MAP Extraction System | Tetracore Inc., Rockville, MD, USA | Spin column | 120 | 2000 | + | + | 4.85 | ||||
Salgado et al., [20] | home-made recipe | centrifugation | 160 | 200 | + | + | + | + | + | ||
MAP DNA SpeedXtract | SpeedXtract Nucleic Acid Kit | Qiagen Lake Constance, Stockach, Germany | inhibitor removal magnetic beads | 25 | 100 | + | + | 4.75 |
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Hansen, S.; Roller, M.; Alslim, L.M.A.; Böhlken-Fascher, S.; Fechner, K.; Czerny, C.-P.; Abd El Wahed, A. Development of Rapid Extraction Method of Mycobacterium avium Subspecies paratuberculosis DNA from Bovine Stool Samples. Diagnostics 2019, 9, 36. https://doi.org/10.3390/diagnostics9020036
Hansen S, Roller M, Alslim LMA, Böhlken-Fascher S, Fechner K, Czerny C-P, Abd El Wahed A. Development of Rapid Extraction Method of Mycobacterium avium Subspecies paratuberculosis DNA from Bovine Stool Samples. Diagnostics. 2019; 9(2):36. https://doi.org/10.3390/diagnostics9020036
Chicago/Turabian StyleHansen, Sören, Marco Roller, Lamia M. A. Alslim, Susanne Böhlken-Fascher, Kim Fechner, Claus-Peter Czerny, and Ahmed Abd El Wahed. 2019. "Development of Rapid Extraction Method of Mycobacterium avium Subspecies paratuberculosis DNA from Bovine Stool Samples" Diagnostics 9, no. 2: 36. https://doi.org/10.3390/diagnostics9020036
APA StyleHansen, S., Roller, M., Alslim, L. M. A., Böhlken-Fascher, S., Fechner, K., Czerny, C. -P., & Abd El Wahed, A. (2019). Development of Rapid Extraction Method of Mycobacterium avium Subspecies paratuberculosis DNA from Bovine Stool Samples. Diagnostics, 9(2), 36. https://doi.org/10.3390/diagnostics9020036