User:Fermenter User/Notebook/BROM A7

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Background color codes:  Pastel Green: Major change of setup this time  yellow: Sung-Hye's correction or question to user

Letter color codes: Black: Basic comment Red: Emergency Information (Power shutdown, building access, Manager's absence etc) Blue: Fermentation Setup information Green: Discussion Orange: Fermentation hrs  Pink: MeOH Induction hrs

BROM A7 (F4)

Day -6 (Jun 12 Fri):
             6/12/2009 (12:00)     Bring BMXY (powder) to fermenter room

Day -4 (Jun 14 Sun):
Building closure 9am - 5pm Power shut-down 10:30 am - 1:30 pm

Day -3 (Jun 15 Mon):
             6/15/2009 (14:00)     Prepare and autoclave additives: Try to make higher concentration, for instance 3. Potassium Phosphate: ?? L                               4. YNB: ?? L                               5. Biotin: ?? L                               6. Glycerol: ?? L                           What is final volume of additives? 7.5 L

            6/15/2009 (12:30)     Autoclave Acid/Base valves, Acid/Base/Antifoam bottles, Water 6/15/2009 (13:58)    Start Fermenter #4 software 6/15/2009 (14:20)    Calculate amount of water = 50L - additives volume (7.5-L) = 42.5L 6/15/2009 (14:30)    Pour BMXY powder and water(42.5 L total) 6/15/2009 (14:30)    Add 10 ml of Antifoam 6/15/2009 (14:48)    Sterilization Routine 6/15/2009 (15:40)    Set rpm at 300 6/15/2009 (15:40)    Purge Air 50 L/min

Day -2 (Jun 16 Tue):
            6/16/2009 (10:00)    Cool down bioreactor at 4°C

             6/16/2009 (00:00)   Take a colony from MD plate in 5 ml of MD. Allow it to gorw at 30°C for 24 hrs.

Day -1 (Jun 17 Wed):
             6/17/2009 (00:00)   Suspend down cells and resuspend in 50 ml BMGY. Allow it to gorw at 30°C for 24 hrs.              6/17/2009 (00:00)   Suspend down cells and resuspend in 2 x 500 ml BMGY. Allow it to gorw at 30°C for 24 hrs.

Day 0 (Jun 18 Thu):
             6/18/2009 (10:00)    Bring additives to fermenter room             6/18/2009 (10:00)     Add Adjutives using pump Picture was taken during transfer of media <Sung-Hye>            6/18/2009 (14:00)     Connect Acid bottle Acid (2000 ml) 6/18/2009 (14:00)    Calibrate dO2 probes 6/18/2009 (14:00)    Start pH control

<Sung-Hye/Raymond>    6/18/2009 (14:45 - 15:40 ) Inoculate cells (O.D: 14.2 Volume: 1000 ml) using pump. <Raymond>             6/18/2009 (15:30)    Fermentation 00:00 No OD check

Day 1 (Jun 19 Fri): <font color=#FF00FF> MeOH Induction Day 0
<Sung-Hye>            6/19/2009 (09:00)    Fermentation 17:30 Connect Base Acid (1850 ml) Base (1600 ml) Autoclave Base bottle and MeOH bottole with connected tube <Sung-Hye/Raymond>    6/19/2009 (11:00) Fermentation 19:30 <font color=#FF00FF> MeOH Induction 00:00 (O.D:22) Contamination? (No) Acid (1850-1800 ml=50 ml) Base (1200-1100 ml=100 ml) Connect MeOH pump <Raymond>             6/19/2009 (17:00) Fermentation 25:30 <font color=#FF00FF> MeOH Induction 06:00 Contamination? (No) Acid (1700 ml Base (1100 ml)                                        Connect MeOH pump

Day 2 (Jun 20 Sat): <font color=#FF00FF> MeOH Induction Day 1
<Raymond>             6/20/2009 (11:00) Fermentation 19:30 <font color=#FF00FF> MeOH Induction 24:00 (O.D:24.29) Contamination? (No) Activity (0.154 /ul) Acid (1600 ml) Base (1100 ml) MeOH (1 ml) <font color=Red> Feed MeOH: Tubing disconnected!! <Sung-Hye>            6/20/2009 (14:00) Fermentation 22:30 <font color=#FF00FF> MeOH Induction 27:00 Autoclave new bottles and tubing for Base and MeOH 6/20/2009 (15:30) Fermentation 24:00 <font color=#FF00FF> MeOH Induction 28:30 Connect Base Connect MeOH (1000 ml) and a pump for <font style=background-color:yellow>continuous feeding: 750 ml/24hr (= 3 times x 0.5%) = 31.25 ml/hr = 0.52 ml/min <font style=background-color:yellow>Use 2rpm Max Watson pump at rpm 25

Day 3 (Jun 21 Sun): <font color=#FF00FF> MeOH Induction Day 2
<Raymond>             6/21/2009 (1:30pm) Fermentation 43:30 <font color=#FF00FF> MeOH Induction 36:00 (O.D:23.15 ) Contamination? (Yes) Activity (0.118/ul) Acid ( 1500 ml) Base (1050 ml) MeOH (400 ml left, then topped up.

Day 4 (Jun 22 Mon): <font color=#FF00FF> MeOH Induction Day 3
<Raymond>             6/22/2009 (2:00pm) Fermentation 00:00 (O.D:24.59 ) Contamination? (Yes) Activity (0.5/5ul from old assay buffer, 1.40/5ul from new assay buffer) Acid (1500ml) Base (1050 ml) MeOH (400ml)

Day 5 (Jun 23 Tue): <font color=#FF00FF> MeOH Induction Day 4
<Raymond>             6/23/2009 (00:00)   Fermentation 00:00 <font color=#FF00FF> MeOH induction 00:00 (O.D:?? ) Contamination? (Yes/No) Activity (?? /ul) Acid (?? ml) Base (?? ml) MeOH (?? ml)

Day 6 (Jun 24 Wed): <font color=#FF00FF> MeOH Induction Day 5
<Raymond>             6/24/2009 (00:00)   Fermentation 00:00 <font color=#FF00FF> MeOH induction 00:00 (O.D:?? ) Contamination? (Yes/No) Activity (?? /ul) Acid (?? ml) Base (?? ml) MeOH (?? ml)


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