Oct 18, 2020
one-step growth curve
- 1South China University of Technology
Protocol Citation: Jiaxin Li 2020. one-step growth curve. protocols.io https://dx.doi.org/10.17504/protocols.io.bnjamcie
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 18, 2020
Last Modified: October 18, 2020
Protocol Integer ID: 43330
Subculture host bacterium in medium of choice plus 2 mM CaCl2 and grow to mid-log phase (ca. 0.5 OD650nm).
Pipette 9.9 mL of the log phage culture into the empty flask and place at the appropriate incubation temperature for 5 min.
Add 0.1 mL of phage preparation to the 9.9 mL culture. (FLASK A)
Transfer 1.0 mL from FLASK A to 9.0 mL of prewarmed medium, mix well. (FLASK B)
Transfer 1.0 mL from FLASK B to 9.0 mL of prewarmed medium, mix well. (FLASK C)
Place FLASK (A, B or C) at 37°C, 180 rpm
Every 10 minutes remove 0.1 mL from the appropriate FLASK (A, B or C) add to the molten OVERLAY; add 0.1 mL of PLATING HOST; mix and pour on surface of UNDERLAY plates. Each experiment should perform in triplicate.
When the overlays have hardened (ca. 15 min) invert the plates and place them in an incubator.
After an appropriate incubation period (ON for E. coli or Pseudomonas aeruginosa) count the plaques on each of the plates.