Biopanning and Phage Amplification

1. E. coli strain K91Kan. The strain was supplied together with the 15-mer library by Prof. G. P. Smith.

2. TBS: 50 mM Tris-HCl, pH 7.4, 50 mM NaCl. Autoclave.

4. TBS/gelatin: 0.1 g gelatin in 100 mL of TBS. Autoclave to dissolve.

6. Elution buffer: 0.1 M glycine-HCl, pH 2.2, 0.1% bovine serum albumin (BSA), 0.01% phenol red.

7. Blocker: 4% BSA in sterile TBS.

8. 80 mM NaCl. Autoclave.

9. NAP buffer: 50 mM NH4H2PO4, pH 7.0 with ammonia, 80 mM NaCl.

10. PEG/NaCl: 10 g of PEG 8000, 11.7 g of NaCl, 47.5 mL of water. Autoclave to dissolve.

11. Stock solutions of antibiotics: 10 mg/mL kanamycin in water; filter-sterilize; 5 mg/mL tetracycline in ethanol; store at -20° C.

12. LB broth: 10 g of bactotryptone, 5 g of yeast extract (both from Difco, Detroit, MI), 5 g of NaCl, and water to 100 mL. Autoclave.

13. LB agar plates: LB broth +16 g/L of bactoagar (Difco, Detroit, MI). Autoclave, allow to cool down to 45-50°C and pour into Petri dishes.

14. LB agar plates +100 Mg/mL kanamycin.

15. LB agar plates + 40 Mg/mL tetracycline + 100 Mg/mL kanamycin. Antibiotics are added after autoclaving, just before pouring into Petri dishes.

16. Plasticware: sterile 35-mm and 90-mm Petri dishes, autoclaved yellow and blue pipet tips, 1.5- and 0.5-mL Eppendorf tubes.

17. Autoclaved toothpicks (antibiotic free).

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