Difference between revisions of "PhiX:Serial dilution"
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==Summary== | ==Summary== | ||
| + | This protocol describes how to prepare serial dilutions in order to enumerate phage from a stock solution. Before you begin you should also prepare for the double agar overlay plaque assay onto which fresh dilutions should be plated. | ||
| − | ''' | + | '''Stock Solution'''<br> |
| − | + | The stock solution has a titer = x PFU/ml (PFU = plaque forming units)<br> | |
| − | + | Alternatively = y pfu in z ul<br> | |
'''SERIAL DILUTION'''<br> | '''SERIAL DILUTION'''<br> | ||
Revision as of 06:49, 20 February 2017
Summary
This protocol describes how to prepare serial dilutions in order to enumerate phage from a stock solution. Before you begin you should also prepare for the double agar overlay plaque assay onto which fresh dilutions should be plated.
Stock Solution
The stock solution has a titer = x PFU/ml (PFU = plaque forming units)
Alternatively = y pfu in z ul
SERIAL DILUTION
Add 100ul (10-1x) phage to 900ul HFB and mix -> tube #1
Transfer 100ul (10-2x) #1 to 900ul HFB and mix -> tube #2
Transfer 100ul (10-3x) #2 to 900ul HFB and mix -> tube #3
and so on...
thoroughly mix between
PLATING
Tube #1 -> plate 100ul (1/10) -> 10-2x
Tube #2 -> plate 100ul (1/10) -> 10-3x
Tube #3 -> plate 100ul (1/10) -> 10-4x
and so on...
MATHS
E.g. count n plaques on plate from 10-7x plate:
x = n * 10-7
OR, if z = 200ul (e.g., mutagenesis)
y = (n * 10-7) * 200/1000
