Team:BYU Provo/Notebook/SmallPhage/Springexp/Period1/Exp/Team:BYU Provo/5.5 Amplification from a plaque test


Small Phage May - June Notebook: Experiments

Small Phage

5.5 Amplification from a plaque test

I) Purpose

Test the procedure for creating phage liquid culture from a single plaque on a plate

II) Expected Outcome

In spot test, we should see plaques until the phage dilution series became too diluted.

III) Reagants Used

E. Coli BL21 overnight from 5.4; ×6 plate from 5.3 T7 phage selection method test; LB from 4.3

IV) Actual Procedure

1. Amplifying phage from a single plaque (5.5)

To a test tube, add 1mL of E coli BL21 overnight (5.4) and 4mL of LB
Stab one of the plaques on a ×6 plate from the 5.3 T7 phage selection method test with a 1mL pipet tip, puncturing the agar. The plaque in question is labeled with a circle.
The pipet tip is then dipped into the overnight/LB solution. The solution is then pipetted up and down several times to maximize the amount of phage that gets into the solution.
The solution was then incubated for approximately 24 hours (5.5 noon - 5.6 2:00pm )

2. Spot test to test amplification results (5.6)

Phage liquid culture was purified and spot test was performed using procedures similar to those in 5.3 T7 phage amplification/purification. Stock created was labeled 5.4
Spot test plates were incubated overnight (5.6 4:00pm - 5.7 12:00pm )

V) Results (5.7)

Our method of amplification from a single plaque was successful. Phage plaque was observed up to -7
5.4 stock spot test.JPG