Commonly Used Media for Phage Growth

Commonly Used Media for Phage Growth

These media will be prepared and may be used for a variety of experiments related to the propagation, titering and genetic crossing of bacterial viruses (bacteriophage). See the protocol Microbiological Media Preparation for principles and practices of media prep. You are to weigh out each reagent listed in bold, add the appropriate amount of distilled water, and dissolve it by heating to 95°C (except for Tryptic Soy Broth which does not need high temp to dissolve because it does not have agar). All media should then be sterilized immediately. The top agar will be melted as needed, aliquoted out in 2 mL portions and kept melted in a 45°C hot block for titering phage.

(Soybean-Casein Digest Medium, USP)

For the cultivation of fastidious and non-fastidious organisms, culturing of coliphage.

Ingredients per liter:
Bacto-Tryptone 17.0 g
Bacto-Soytone 3.0 g
Bacto-Dextrose 2.5 g
Sodium Chloride 5.0 g
Dipotassium Phosphate 2.5 g
TRYPTONE AGAR (For growth of phage)* grams/600 mL:

(Make 3 bottles for a class of 20)

tryptone                                     6.0 g
NaCl                                         0.5 g
1 M CaCl2 (2 mL/L medium)     1.2 mL
agar                                           6.6 g

For use in titering phage, and preparing a poured a lawn of bacteria in top agar.

grams/200 mL:

tryptone (1%)                          2.0 g
KCl (0.5%)                             1.0 g
agar (0.6%)                             1.2 g

Mix, distribute 2 mL/13 x 100 mm test tube, cap and sterilize.

Before use, melt in 100°C water, cool to 45°C. (Alternately, sterilize in bottle, melt just before use, aliquot out into 45°C capped sterile Wasserman tubes.)
LB AGAR, MILLER (Luria-Bertani) 24 g/600 mL

For the cultivation Escherichia coli, titering of coliphage, especially the fastidious lambda bacteriophage.

Ingredients per liter:
Bacto-Tryptone                   10.0 g
Bacto-Yeast Extract              5.0 g
Sodium Chloride                  10.0 g
Bacto-Agar                         15.0 g

* From Bob Kirchen at Carolina Biological, 2/7/95

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