IGEM:UC-Merced/2009/Notebook/E. hydro Express
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Beginning Tuesday November 13, 2012 | Beginning Tuesday November 13, 2012 | ||
| - | + | 11/13/2012 | |
Generation P1 Phage Stock | Generation P1 Phage Stock | ||
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25000 ug/ml (V1) = 250 ug | 25000 ug/ml (V1) = 250 ug | ||
V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1 | V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1 | ||
| - | Incubate kanomycin in LB broth until 11am | + | Incubate kanomycin in LB broth until 11am 11/14/12 |
| - | + | 11/14/12 | |
P1 Phage | P1 Phage | ||
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25000 ug/ml (V1) = 250 ug | 25000 ug/ml (V1) = 250 ug | ||
V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1 | V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1 | ||
| - | Incubate kanomycin in LB broth until 10:30 am 10/15/12 | + | Incubate kanomycin in LB broth until 10:30 am 11/15/12 |
| + | |||
| + | 11/15/12 | ||
| + | |||
| + | 1. JW1228-1 overnight culture grew. The next step in the procedure was carried out according to Yale University. | ||
| + | 2. Preparation of 10ml containing 20% Glucose solution | ||
| + | a. Calculations | ||
| + | 20g/100 ml = x/10ml | ||
| + | x=2 g of glucose | ||
| + | 3. The 20% glucose solution was made using filtered water and 2 g of glucose. The resulting 10 ml of glucose solution was filtered. | ||
| + | 4. Preparation of Overnight JW1228-1 culture to be used in P1 phage infection | ||
| + | a. Calculation to make a 1:100 dilution of JW1228-1 overnight culture | ||
| + | 1/100=x/5ml | ||
| + | 100x=5ml | ||
| + | x=0.05 ml or 50 ul | ||
| + | b. Two culture tubes were created for P1 phage infection | ||
| + | i. Each tube include: 50 ul of overnight culture, 50 ul of 20% glucose, and 25 ml of 1M CaCl2 | ||
| + | 5. JW1228-1 overnight culture prepared for P1 phage solution to be incubated until 11:55 on 10/15/12 | ||
Revision as of 15:29, 15 November 2012
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Description/Abstract
Notes
Lab Notebook Beginning Tuesday November 13, 2012 11/13/2012 Generation P1 Phage Stock 1. Prepared P1 Stock from Yale University
a. LB
b. Kanamycin stock 25mg/ml to add to strain JW1228-1
C1V1=C2V2
25000 ug/ml (V1) = 50ug/ml (5ml)
C1 = 25000 ug/ml --> kanamycin concentration
C2 = 50 ug/ml
V2 = 5 ml broth
25000 ug/ml (V1) = 250 ug
V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1
Incubate kanomycin in LB broth until 11am 11/14/12
P1 Phage 1. Overnight culture did not grow. Possible reason is: The JW1228-1 culture taken from the 4°C refrigerator was old and was probably no longer living. To address this possibility we will redo the procedure using frozen JW1228-1 stock. 2. Prepared new overnight culture with frozen JW1228-1 glycerol stock (following the procedure carried out on 10/13/12) a. LB
b. Kanamycin stock 25mg/ml to add to strain JW1228-1
C1V1=C2V2
25000 ug/ml (V1) = 50ug/ml (5ml)
C1 = 25000 ug/ml --> kanamycin concentration
C2 = 50 ug/ml
V2 = 5 ml broth
25000 ug/ml (V1) = 250 ug
V1 = 0.01 ml = 10 uL --> kanamycin to add to strain JW1228-1
Incubate kanomycin in LB broth until 10:30 am 11/15/12
11/15/12 1. JW1228-1 overnight culture grew. The next step in the procedure was carried out according to Yale University. 2. Preparation of 10ml containing 20% Glucose solution a. Calculations
20g/100 ml = x/10ml
x=2 g of glucose
3. The 20% glucose solution was made using filtered water and 2 g of glucose. The resulting 10 ml of glucose solution was filtered. 4. Preparation of Overnight JW1228-1 culture to be used in P1 phage infection a. Calculation to make a 1:100 dilution of JW1228-1 overnight culture
1/100=x/5ml
100x=5ml
x=0.05 ml or 50 ul
b. Two culture tubes were created for P1 phage infection
i. Each tube include: 50 ul of overnight culture, 50 ul of 20% glucose, and 25 ml of 1M CaCl2
5. JW1228-1 overnight culture prepared for P1 phage solution to be incubated until 11:55 on 10/15/12 |
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