1.) you read an overnight culture of E. coli and its OD600 is 0.6. how many CFUs
ID: 201472 • Letter: 1
Question
1.) you read an overnight culture of E. coli and its OD600 is 0.6. how many CFUs/mL are in this culture
2.) you need to inoculate 100 microliters of fresh, sterile LB to reach 10^3 CFU/mL from an overnight E. coli culture above. what volume of your overnight culture will you need to use in your inoculation?
3.) how would you accuratley deliver the volume you determined in #2 above, using a p1000, p200, p20 within its pipetting limits and other normal labware?
Explanation / Answer
1) Given to the distortion in the picture, i have best assumed the CFU/ml to be 6 * 106.
2) S1V1=S2V2 , this formula can work for all dilutions, where S is concentration and V is volume, before and after dilution, So,
6*106 * V1 = 103 * 100; V1= 0.016 ml = 16 microlitres
3) The limits are in microlitres( 1 ml = 1000 microlitres)
P20: 2.0 - 20.0 , can be done easily by fixing the volume limit of the pipette to be 16 microlitres
P200: 20 - 200,
S1V1=S2V2
6* 106 * 20 = 103 * V2
V2= 120000 microlitres= 120 ml, so we inoculate in 120 ml, and after mixing we discard 20 ml. Alternatively, we can do serial dilution(more preferable) shown for next
P1000: 200 - 1000.
S1V1=S2V2
6* 106 * 200 = 105 * V2
V2= 12000 microlitres= 12 ml, so we inoculate first 200 microlitres 12 ml which will have a cfu of 105, and then
S1V1=S2V2
105 * V1 = 103 * 100 ml
V1= 1 ml, so we inoculate first 1 ml making the volume upto 100 which will now have a cfu of 103
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