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Polymerase Chain Reaction (PCR) Amplification Protocol_
| Pre-denaturation | Denaturation | Annealing | Extension | Final extension | Number of cycles |
|---|---|---|---|---|---|
| 95°C, 5min | 95°C, 30s | 58°C, 30s | 72°C, 1min 30s | 72°C, 7min | 35 |
Identification results of 17 bacterial strains_
| Strain No. | Scientific name | Query coverage | Identities | Sequence ID |
|---|---|---|---|---|
| CHF-2 | Streptomyces heliomycini | 100% | 99.22% | NR_041197.1 |
| CHF-3 | Streptomyces plumbiresistens | 100% | 99.21% | NR_044518.1 |
| CHF-5 | Streptomyces alfalfae | 100% | 98.79% | NR_147713.1 |
| CHF-6 | Streptomyces huasconensis | 100% | 100.00% | NR_178969.1 |
| CHF-7 | Streptomyces spiroverticillatus | 100% | 99.50% | NR_112582.1 |
| CHF-8 | Niallia nealsonii | 100% | 99.22% | NR_044546.1 |
| CHF-9 | Streptomyces alfalfae | 100% | 98.71% | NR_147713.1 |
| CHF-10 | Streptomyces kanamyceticus | 99% | 98.88% | NR_043822.1 |
| CHF-11 | Acinetobacter seifertii | 100% | 100.00% | NR_134684.1 |
| CHF-12 | Streptomyces alfalfae | 98% | 97.99% | NR_147713.1 |
| CHF-14 | Peribacillus frigoritolerans | 100% | 99.93% | NR_117474.1 |
| CHF-15 | Streptomyces alboniger | 100% | 99.35% | NR_043228.2 |
| CHF-16 | Streptomyces alfalfae | 100% | 97.59% | NR_147713.1 |
| CHF-17 | Streptomyces spiroverticillatus | 100% | 99.43% | NR_112582.1 |
| CHF-18 | Streptomyces spiroverticillatus | 100% | 99.50% | NR_112582.1 |
| CHF-19 | Pseudarthrobacter siccitolerans | 100% | 99.50% | NR_108849.1 |
| CHF-20 | Streptomyces aureus | 99% | 99.06% | NR_025663.1 |
Results of inhibition zone determination of antibacterial activity of strains_
| Strain No. | Bacteriostatic diameter/cm | ||
|---|---|---|---|
| Staphylococcus aureus | Proteus vulgaris | Candida albicans | |
| CHF-2 | – | – | – |
| CHF-3 | 1.52 | 1.56 | 1.55 |
| CHF-5 | 4.25 | – | – |
| CHF-6 | 2.27 | – | 2.52 |
| CHF-7 | 1.49 | – | – |
| CHF-8 | – | – | – |
| CHF-9 | 1.78 | 1.55 | – |
| CHF-10 | 2.11 | – | – |
| CHF-11 | 1.62 | 1.53 | 1.54 |
| CHF-12 | 3.34 | 3.35 | – |
| CHF-14 | 2.14 | 1.50 | 1.53 |
| CHF-15 | 1.20 | 2.73 | – |
| CHF-16 | 4.14 | 2.37 | – |
| CHF-17 | 1.49 | – | 1.72 |
| CHF-18 | 1.47 | 1.37 | 1.69 |
| CHF-19 | – | – | – |
| CHF-20 | 2.20 | – | – |
Salt tolerance of 17 bacterial strains_
| Strain No. | NaCl concentration (g/l) | |||
|---|---|---|---|---|
| 25 | 50 | 75 | 100 | |
| CHF-2 | + | + | + | – |
| CHF-3 | + | – | – | – |
| CHF-5 | + | – | – | – |
| CHF-6 | + | + | + | – |
| CHF-7 | + | – | + | – |
| CHF-8 | + | – | – | – |
| CHF-9 | + | – | – | – |
| CHF-10 | + | – | – | – |
| CHF-11 | + | + | + | + |
| CHF-12 | + | – | – | – |
| CHF-14 | + | + | + | – |
| CHF-15 | + | + | – | + |
| CHF-16 | + | – | – | – |
| CHF-17 | + | + | – | – |
| CHF-18 | + | – | + | + |
| CHF-19 | + | – | + | – |
| CHF-20 | + | + | – | – |
Primer design_
| Primer name | Sequence |
|---|---|
| 27F | 5′-AGAGTTTGATCCTGGCTCAG-3′ |
| 1492R | 5′-GGTTACCTTGTTACGACTT-3′ |
PCR Amplification Reaction System_
| Reagent | Volume |
|---|---|
| Genomic DNA (20 ng/μl) | 1.0 μl |
| 10× Buffer (containing 2.5 mM Mg2+) | 5.0 μl |
| Taq Polymerase (5 U/μl) | 1.0 μl |
| dNTP (10 mM) | 1.0 μl |
| 27F Primer (10 μM) | 1.5 μl |
| 1492R Primer (10 μM) | 1.5 μl |
| ddH2O | 39.0 μl |
| Total volume | 50.0 μl |
Alkali resistance of 17 bacterial strains_
| Strain No. | pH | |||
|---|---|---|---|---|
| 8 | 9 | 10 | 11 | |
| CHF-2 | + | + | + | + |
| CHF-3 | + | + | + | + |
| CHF-5 | + | + | + | + |
| CHF-6 | + | + | + | + |
| CHF-7 | + | + | + | + |
| CHF-8 | + | + | + | – |
| CHF-9 | + | + | + | + |
| CHF-10 | + | + | + | + |
| CHF-11 | + | + | + | + |
| CHF-12 | + | + | + | + |
| CHF-14 | + | + | + | + |
| CHF-15 | + | + | + | + |
| CHF-16 | + | + | + | + |
| CHF-17 | + | + | + | + |
| CHF-18 | + | – | – | + |
| CHF-19 | + | – | – | – |
| CHF-20 | + | + | + | + |