Genetics - Transduction with bacteriophage P1

Jeff Lawrence (G12)

1. Grow recipient cells overnight in LB at 37°C with shaking.

2. Dilute add 100 µL cells to 5 mL LB. Grow at 37°C with shaking for 60 min, or until density reaches Klett 50, (OD650 = 0.3).

3. Centrifuge cells for 8 min at 6000 rpm. Discard supernatant.

4. Resuspend cells in 2.5 mL (1/2 volume) of P1 Dilution Buffer:

	5 mM	CaCl2	500 µL	1 M CaCl2
	10 mM	MgSO4	1 mL	1 M MgSO4
			99 mL	H₂O

5. Chill cells on ice for 30 - 45 min.

6. Mix 100 µL cells with 10 µL, 50 µL, and 100 µL phage in three separate tubes.

7. Incubate at 30°C for 30 min.

8. Add 150 µL 1M NaCitrate to each tube.

9. Plate on selective medium.

10. Incubate at 37°C overnight.

Last Update: Thursday June 19 2014

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Eric Kofoid eckofoid at ucdavis.edu