Serial Dilution Problems
1. You are planning an experiment for which you need the exact concentration of an E coli. Culture. For this purpose you grow E coli. Overnight in LB broth (this is your stock solution). You prepare a 1:5 dilution of your stock solution and measure the OD which is 0.7. Since you want to make sure that your OD reading corresponds with number of viable cells you do the following. First you do a 1:1000 dilution of your stock (this is your first dilution). Then you perform a 1:100 dilution (this is your second dilution). Finally you plate 10 µL of dilution #2 onto an LB plate and incubate at 37°C. The next day you can count 4 colonies on your plate. Is this the results you expected?
A) No, I would expect 10 times more colonies. Maybe the plate I used to plate the cells was not an LB agar but a GMS plate
B) No, I would expect 2 times more colonies because the concentration of my stock is 7 x 108 cells/mL
C) No, I would expect 10 times more colonies, because the total dilution factor is 1:106
D) No, I would expect 10 times more colonies. Maybe I used the 1:5 dilution instead of the stock to prepare dilution 1.
E) None of the above
2. A student completes a serial dilution from an E coli culture grown in LB. She uses a 100 fold dilution factor and completes 3 dilutions. From her third dilution she plates 100 µL. She then incubates the plate for 24 hours at 37°C. The student finds 47 colonies on her plate. What was the true concentration of the stock culture?
3. You are given an Escherichia coli culture that has 2500 cells/ml. You plate 10µL of this culture on an LB plate. How many colonies will you see on the LB plate after incubation at 37°C for 24 hours?
4. You have 0.72ml of LB in an Eppendorf tube and transfer 80µl into the tube. What is your dilution factor?
5. You grow a culture of E. coli to an OD reading of 0.8. Your instructor wants 40colonies on a plate. Ideally, you want to only spread 100ul on a LB plate. How would you carry out the proper dilutions in order to arrive at 40 cells on a plate? Please show your calculations and all work. (1 O.D. = 1 x 108 cells/mL).
6. You grow a culture of E. coli to an OD reading of 0.9. Your instructor wants 300 colonies on a plate. Ideally, you want to only spread 100ul on a LB plate. How would you carry out the proper dilutions in order to arrive at 300 cells on a plate? Please show your calculations and all work. (1 O.D. = 1 x 108 cells/mL).
7. You grow a culture of E. coli and when taking the OD reading the spectrophotometer gives you an OD value above 1. You decide to do a ten-fold dilution of your culture and take the OD reading of the diluted culture. The OD reading of the diluted culture is 0.5. Your instructor wants 500 colonies on a plate. Ideally, you want to only spread 50ul on a LB plate. How would you carry out the proper dilutions in order to arrive at 500 cells on a plate? Please show your calculations and all work. (1 O.D. = 1 x 108 Cells/mL)