Bacterial 16S rDNA Amplification Protocols
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Corresponding Organization :
Other organizations : University of Vienna, Max Planck Institute for Marine Microbiology, University of Bremen, Constructor University
Protocol cited in 261 other protocols
Variable analysis
- Primer pair (i): S-D-Bact-0341-b-S-17 and S-D-Bact-0785-a-A-21
- Primer pair (ii): S-D-Bact-0008-a-S-16 and S-D-Bact-0907-a-A-20
- Quantity and quality of the extracted DNA
- Reaction volume: 50 µl
- BSA concentration: 0.3 mg/ml
- DTNPs concentration: 250 µM
- Primer concentration: 0.5 µM for each primer
- DNA Polymerase: 0.02 U Phusion High-Fidelity DNA Polymerase
- Buffer: 5x Phusion HF Buffer containing 1.5 mM MgCl2
- PCR conditions: initial denaturation at 95°C for 5 min, 25 cycles of denaturation (95°C for 40 s), annealing (2 min), and extension (72°C for 1 min), and a final extension step at 72°C for 7 min
- Annealing temperature for primer pair (i): 55°C
- Annealing temperature for primer pair (ii): 44°C
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