Preparing a Soil Bacterial Culture for Bacteriophage Infection

Abstract

Source: Cross, T., et al., An Optimized Enrichment Technique for the Isolation of Arthrobacter Bacteriophage Species from Soil Sample Isolates. J. Vis. Exp.(2015)

This video demonstrates the preparation of a bacterial culture for bacteriophage production. It covers the steps of inoculating a soil bacterium into a baffled flask, promoting uniform growth through aeration and shaking, and monitoring cell density using optical density measurements. Once the culture reaches the optimal growth phase, it is ready for efficient bacteriophage production.

Protocol

Preparation of Arthrobacter Cells for Phage Isolation

• Culture Arthrobacter sp. KY3901 colonies streaked on an Luria Bertani (LB) agar plate incubated at 30 °C for 2-3 days. Pick a colony and use a sterile loop to add it to 250 ml of LB broth in a baffled culture flask and incubate in a shaking incubator at 225 rpm at 30 °C.
• Allow approximately 24 hr of growth to obtain late exponential/early stationary phase cells for phage infection experiments. Monitor the state of bacterial growth closely to prevent cells from entering the middle to late stationary growth state.
  NOTE: Cell growth should consist of an optical density OD₆₀₀ between 0.5-.0.7 for the phage isolation and purification procedures/steps.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
LB Broth powder	Fisher	BP9722-2
Granulated Agar	Fisher	BP1423-2
Eppendorf Tubes	Fisher	05-408-129
5 ml pipets individual	Fisher	13-678-11D
50 ml conical tubes	Fisher	76002844
15 ml conical tubes	Fisher	76002845
10 ml pipets individual	Fisher	13-676-10J
25 ml pipets individual	Fisher	13-676-10K