1. A grad student has a culture of E. coli with a total volume of 1,800 mL, and OD reading at 550 nm of 1.7. He transfers 1 mL culture into 9 mL tryptone broth. Next, he transfers 0.1 mL of this dilution into 0.9 mL tryptone broth, and repeats this two more times to make a total of 4 serial dilutions. He then pipets 200 uL from the fourth dilution on an agar plate, spreads it over the surface, and puts the plate in an incubator to grow overnight. The next day, he counts 386 cfus (colony forming units) on the plate.
a. What is the dilution in the third tube? Answer with the exponent to the power of 10, that is ? in 10? b. What is the dilution of cells spread onto the agar plate? Answer in decimal notation.
c. How many viable cells are present in the 1800 mL starting culture? Answer in decimal notation.
1. A grad student has a culture of E. coli with a total volume of 1,800 mL, and OD reading at 550 nm of 1.7. He transfers 1 mL culture into 9 mL tryptone broth. Next, he transfers 0.1 mL of this dilution into 0.9 mL tryptone broth, and repeats this two more times to make a total of 4 serial dilutions. He then pipets 200 uL from the fourth dilution on an agar plate, spreads it over the surface, and puts the plate in an incubator to grow overnight. The next day, he counts 386 cfus (colony forming units) on the plate.
a. What is the dilution in the third tube? Answer with the exponent to the power of 10, that is ? in 10? b. What is the dilution of cells spread onto the agar plate? Answer in decimal notation.
c. How many viable cells are present in the 1800 mL starting culture? Answer in decimal notation.
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A student grows a culture of E. coli in 50 ml of liquid medium contained in a 250 ml Erlenmeyer flask and decides to pour plate samples of the culture onto LB agar plates. (LB stands for Luria Broth. Itâs simply a type of broth on which E. coli grow well.) Before plating, the student sets up several dilutions of the culture (indicated in the table) and plates 0.2 ml of each by pour plating. S/He does this in duplicate for each dilution. S/He incubates all of her LB agar plates in a 37 ºC incubator and counts the colonies on each plate later. Her/his results are as follows
| Total dilution factor | Plate A | Plate B |
| 10-6 | 756 | 349 |
| 10-7 | 73 | 76 |
| 10-8 | 7 | 8 |
a) Determine the average density in cfu/ml of the culture based on the relevant results. Give your answer in scientific notation with three significant figures.
b) Significantly fewer colonies grew on Plate B at the 10-6 dilution as compared to Plate A at the same dilution. Which plate (Plate A or Plate B) is more likely to be a true indication of the number cells that were present in the culture at the 10-6 dilution? Explain your reasoning.
FOR #4 A & B I don't know if I did it correctly.....Please check & the other questions I'm having a heard time on. Thanks!
A. You have been given a tube of E. coli. You are asked to make 1 mL total volume of 10-1 dilution of the bacterial culture. Explain how you would do this. Show all necessary calculations.
____ ml cells + _____ ml water = 1 ml (total volume)
V1 D1 X V2 D2
1 mL X 1 = V2 X 10^-1
V2 = 1 mL/10^-1 so V2 = 10 mL.
But because V1 is part of V2. The answer is: 10 mL-1 mL= 9 mL of diluent must be added.
V1 D1 X V2 D2
1 mL X 1 = V2 X 10^-2
V2 = 1 mL/10^-2 so V2 = 100 mL.
But because V1 is part of V2. The answer is: 100 mL-1 mL= 99 mL of diluent must be added.
You have bacteria at a concentration of 5 x 108 CFU/mL. You spread 1 mL of this sample on an agar plate to obtain isolated colonies. How many colonies do you expect to find the next day after incubation at 370C? Can you count these colonies? Can you use this plate for determining concentration?
A. You have bacteria at a concentration of 1 x 103 CFU/mL (in real life â you donât know this, but we are just working on math skills here). You transfer 1 mL of this sample into 9 mL of water and then spread 1 mL on a plate of agar. How many colonies do you expect to find the next day after incubation at 370C? Can you count these colonies? Can you use this plate for determining concentration?
You take 0.05 mL of a culture of bacteria at a concentration of 4 x 107 CFU/mL, and add 4.95 mL of water to it. What is the dilution that you have performed? What is the concentration of bacteria (CFU/mL) in the diluted culture?
A. You have diluted a sample by 1000 fold (1/1000) and plated 1 mL on an agar plate. You observe 55 colonies. What was the concentration of the original sample in CFU/mL?
A. A bacterial sample has a concentration of 3 x107CFU/mL. You make serial dilutions of 10-3 followed by 10-2 and 10-1 dilutions. You finally plate 1 mL of the last dilution on an agar plate and incubate it at 370C. What is the total dilution? How many colonies do you expect to see on the plate?
Total dilution:
# of colonies expected on plate:
C. This time, you see 10 times fewer colonies than you had expected to see.What could have gone wrong? How will you fix this problem?
A bacterial sample has a concentration of 2 x106 CFU/mL. Show a scheme of dilutions to obtain 30-300 colonies on a plate. Your scheme should contain the volume of diluent (sterile water), volume of sample transferred each time, the concentration of bacteria (CFU/mL) in each dilution. You can assume you are plating 1 mL of the dilution on plates of nutrient agar. See Figure 1 in protocol.
These are the results of the experiment described in the protocol. From the data shown in the results section of Standard Plate Count protocol, calculate the concentration of bacteria (CFU/mL) in the original sample of E. coli. Show your calculations. Hint: remember that we donât use plates that have less than 30 or more than 300 colonies to do these calculations.
