User:Fermenter User/Notebook/MACG E6

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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 Purple: Induction hrs

MACG E6

Day 0 (Mar 17 Tue)
 pH sensor: monitor or control pH   dO2 sensor: monitor or control dO2 Condenser: Gas outlet OD probe: monitor cell growth ==> Not used! Replate it with septum Triple: Addition of media, Acid or Base if necessary MeOH sensor: Monitor EtOH (byproduct)  MeOH sensors also detects ethanol! :See reference #6]            3/17/2009 (10:00)  Pour media 3/17/2009 (10:30) pH calibration 3/17/2009 (11:10) Autoclave bioreactor ==> 3 bioreactors 3 empty bottles/fermenter Carbon source, Inoculum, Feeding bottle 3/17/2009 (13:30) Start Temp Control 3/17/2009 (13:40) Connect MeOH sensor 3/17/2009 (13:42) Add antifoam 0.5 ml/fermenter 3/17/2009 (13:46) Start F1 software 3/17/2009 (13:47) Start F2 software 3/17/2009 (13:48) Start F3 software 3/17/2009 (14:00) Add Carbon source using pump at 30°C 3/17/2009 (14:05) Connect Acid (250 ml) and Base (400 ml) for Fer#2 and Fer#3 3/17/2009 (14:05) Set pH for Fer#2 and Fer#3 3/17/2009 (14:07)  Calibrate dO2 3/17/2009 (14:07)  Set dO2   3/17/2009 (14:45) Fermentation 00:00 Inoculation 30 ml each using a pump.                no readings taken today

Day 1 (Mar 18 Wed)
           3/18/2009 (7:00) F2: Acid (210 ml) Base (350 ml) F3: Acid (250 ml) Base (370 ml)                3/18/2009 (14:00)  Fermentation 23:15 F1: OD (63.7) Contamination? (N)Pink, budding F2: OD (60.4) Contamination? (N)Yellowish pink, not many buds F3: OD (78.6) Contamination? (N)Pink budding.            3/18/2009 (15:30)  Fermentation 24:45 F3: Change set point of pH                   3/18/2009 (17:00)  Fermentation 26:15 F1: OD (64.5) Contamination? (N) F2: OD (57.0) Contamination? (N) F3: OD (76.5) Contamination? (N)

Day 2 (Mar 19 Thu)
           3/19/2009 (07:00) F2: Acid (150 ml) Base (270 ml) F3: Acid (250 ml) Base (110 ml)                3/19/2009 (12:00)  Fermentation 45:15 Induction 00:00 F1: OD (73.6) Contamination? (N) pink, approx 1% budding F1:Induction 00:00 F2: OD (61.4) Contamination? (N) paler pink, no budding F2:Induction 00:00 F3: OD (59.1) Contamination? (N) pink, approx 1% budding, -probably due to larger volume F3:Induction 00:00 <Sung-Hye and Isis>  3/19/2009 (12:00)  Fermentation 45:15 Pepstatin added (12:30) diluted F1: Connect Pump to 500 ml of Feeding media F1:Induction 00:00 Watson pump: 9 rpm (10rpm = 12.8 ml/h) <Sung-Hye and Isis>  3/19/2009 (12:00)  Fermentation 45:15 Pepstatin added (12:30) diluted F2: Connect Pump to 500 ml of Feeding media F2:Induction 00:00 Watson pump: 9 rpm (10rpm = 12.8 ml/h) <Sung-Hye and Isis>  3/19/2009 (12:00)  Fermentation 45:15 Decanted 300 ml                             Pepstatin added (12:30) diluted F3: Connect Pump to 1000 ml (x2 diluted) of Feeding media F3:Induction 00:00 Watson pump: 1 rpm (32 rpm pump used)



Day 3 (Mar 20 Fri)
<Sung-Hye>           3/20/2009 (07:00) F2: Acid (130 ml) Base (210 ml) F3: Acid (0 ml) Base (0 ml) Transfer 50-ml of Acid and Base from F2 to F3   <Isis>                3/20/2009 (08:00) Fermentation 65:15 F1: OD (119.6) F1:Induction 20:00 Contamination?(N)no budding, multiple vacuoles, pinkest cells, SN most yellow, smells bad F2: OD (98.7) F3:Induction 20:00 Contamination? (N) approx 5% budding, pink cells, yellow SN, isoamyl acetate, filter chnaged F3: OD (73.1) F2:Induction 20:00 Contamination? (N) approx 5% budding, least pink cells, least yellow media, most isoamyl acetate smell. ---(volume normalization)-- F1: 1-L (media+cells) + Feeding (250 ml) = 1.25-L F2: 1-L (media+cells) + Acid (120 ml) + Base (190 ml) + Feeding (230 ml) = 1.54-L F3: 1-L (media+cells) + Acid (250 ml) + Base (400 ml) + Feeding (580 ml) = 2.23-L ---     <Isis>                3/20/2009 (10:00) Fermentation 67:15 F1: OD (134.3)Contamination? (N) Harvest F1:Induction 22:00 F2: OD (86.2) Contamination? (N) F2:Induction 22:00 F3: OD (73.9) Contamination? (N) F3:Induction 22:00 <Isis>               3/20/2009 (12:00) Fermentation 69:15 F2: OD (??) Contamination? (N) F2:Induction 24:00 F3: OD (??) Contamination? (N) F3:Induction 24:00 <Isis>               3/20/2009 (14:00) Fermentation 71:15 F2: OD (94.4) Contamination? (N) F2:Induction 26:00 F3: OD (65.2) Contamination? (N) F3:Induction 26:00 <Isis>               3/20/2009 (16:00) Fermentation 73:15 F2: OD (98.7) Contamination? (N) F2:Induction 28:00 F3: OD (78.5) Contamination? (N) F3:Induction 28:00 <Isis>               3/20/2009 (18:00) Fermentation 75:15 F2: OD (??) Contamination? (N) F2:Induction 30:00 F3: OD (??) Contamination? (N) F3:Induction 30:00