Question 2 . You are given a yeast culture of concentration 1.8 x 10 10 cfu/ml.

Question

Question 2. You are given a yeast culture of concentration 1.8 x 1010 cfu/ml. You are asked to make a 1.8 x 10-8 dilution from this stock culture. The volumes in your tube(s) should not exceed 10 ml and the volumes you pipette should not be less than 0.1 ml.

(i) What is the CDF for this dilution?

(ii) Given your volume constraints, can you do this dilution in a single step? If yes, show me how. If no, answer questions (iii) and (iv) below.

(iii) If no, demonstrate (draw or grid out) how you would set up the dilution. For each step, indicate in an orderly fashion the following information: V2, V1, diluent volume, DF for each dilution step, CDF up to that step, yeast concentration at that step.

(iv) Is this a serial dilution or a dilution series?

(v) What volume should you plate from the 1.8 x 10-8 dilution to get 81 colonies

Explanation / Answer

Given a yeast culture of concentration 1.8 x 1010 cfu/ml.

To prepare = 1.8 x 10-8 dilution from this stock culture.

Final volume of the tube = 10 ml

N1V1 (stock) = N2 V2 (working solution)

1.8 x 1010 * V1 = 1.8 x 10-8 * 10

V1 = 10-17 ml (To solve this in one step, pipette out 10-17ml stock solution and mix it in 10ml distilled water. This combination is very difficult to attempt, and will result in a lot of manual errors.)

Also, it is given, that Volumes you pipette should not be less than 0.1 ml; therefore serial dilutions should be used.

Step 1:-Take out 0.1 ml stock solution and mix it in 10 ml diluent i.e distilled water = 0.1/10.1= i.e we make 1/ 101 dilution; diluted 101 times.

Step 2:-Again take 0.1 ml of this dilution (of step 1) and mix it in 10 ml distilled water = 0.1/ 10.1 i.e. we again dilute it 101 times ; CDF , cumulative dilution factor is 101*101= 10201 times

Step 3:- In this repeat step 2 , CDF comes out to be 10201 * 101 = 1030301 times

Step 4 :- Repeat this 8 times to get 10-8 dilution = (101)8 will be the final CDF.

CFU/ml = no. of colonies / volume plated

1.8 * 10-8 = 81 / V

V = 81/ 1.8 * 10-8

V = 45 * 108 ml = 4.5 * 109ml should be plated to get 81 colonies.

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