User:Fermenter User/Notebook/MACG E5

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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 E5 (F1 and F2)

Day -1 (Feb 23 Mon)
 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]            2/23/2009 (9:30)  Pour media 2/23/2009 (9:30) pH calibration pH sensor #1 (F1) pH sensor #2 (F2) pH sensor #3 (F3) 2/23/2009 (9:30) Antifoam added 2/23/2009 (10:15) Autoclave bioreactor ==> 3 bioreactors 3 empty bottles/fermenter Carbon source, Inoculum, Feeding bottle 2/23/2009 (12:00) Start Temp Control 2/23/2009 (13:30) Add Carbon source using pump at 30°C 2/23/2009 (13:39) Start F1 software 2/23/2009 (13:40) Start F2 software

Day 0 (Feb 24 Tue)
Sung-Hye Class 9:30-11:00

           2/24/2009 (08:00)   Connect MeOH sensor 2/24/2009 (08:00)  Calibrate dO2 2/24/2009 (08:00)  Set dO2   2/24/2009 (12:00) Fermentation 00:00 Inoculation 30 ml each using a pump.                no readings taken today

Day 1 (Feb 25 Wed)
               2/25/2009 (9:00)  Fermentation 21:00 F1: OD (58.5) Contamination? (N)smaller, rounder cells, very clumpy F2: OD (49.3) Contamination? (N)larger oval cells, budding well, pink color developing already.                2/25/2009 (10:00)  Fermentation 22:00 F1: OD (61.5) Contamination? (N) F2: OD (57.0) Contamination? (N)                2/25/2009 (11:00)  Fermentation 23:00 F1: OD (64.3) Contamination? (N) F2: OD (59.7) Contamination? (N)                2/25/2009 (12:00)  Fermentation 24:00 F1: OD (63.5) Contamination? (N) F2: OD (59.2) Contamination? (N) Induction of F1 (Transfer to F3)   2/25/2009 (13:40) Fermentation 25:40 <font style=background-color:yellow>Spin-down cell pellet and transfer to second bioreactor (13:48) Start software for F3    <Sung-Hye and Isis>   2/25/2009 (14:00)  Fermentation 26:00 F3: Transfer cells + Carbon source (5ml of 40% Gal) to second bioreactor by pump Pepstatin and Benzamidine.HCl (1 mM Final) were added via septum F3: OD (53.2) Contamination? (N) <Sung-Hye and Isis>  2/25/2009 (15:00) Fermentation 27:00 F3: Connect Pump to ?? ml of Feeding media F3:Induction 00:00 Watson pump: 9 rpm (10rpm = 12.8 ml/h) Induction of F2    <Sung-Hye and Isis>   2/25/2009 (16:00) Fermentation 28:00 <font style=background-color:yellow>Spin-down cell pellet and pour back to same bioreactor F2: OD (59.0) Contamination? (N) <Sung-Hye and Isis>  2/25/2009 (16:00)  Fermentation 28:00 F2: Transfer cells + Carbon source (5ml of 40% Gal) to second bioreactor by syringe Pepstatin and Benzamidine.HCl (1 mM Final) were added via septum <Sung-Hye and Isis>  2/25/2009 (17:00) Fermentation 29:00 F2: Connect Pump to ?? ml of Feeding media F2:Induction 00:00 Watson pump: 9 rpm (10rpm = 12.8 ml/h) -



Day 2 (Feb 26 Thu)
Sung-Hye Class 9:30 - 11:00

<Isis>               2/26/2009 (07:30)  Fermentation 55:30 F3: OD (96.6) Contamination? (N) F3:Induction 16:30 Cells budding, small and round, clumping. Cell pellet cream/white. Smell from fermenter sweet like banana flavored milk. <font style=background-color:yellow> --> See Reference #9, #10 about "Banada Odor": It is [Isoamyl Acetate] F2: OD (118.8) Contamination? (N) F2:Induction 14:30 Cells budding, larger, vacuoles present (or multiple buds). Cell pellet pink. <Isis>               2/26/2009 (09:00)  Fermentation 57:00 F3: OD (-) Contamination? (N) Fe:Induction 18:00 F2: OD (-) Contamination? (N) F2:Induction 16:00 <Isis>               2/26/2009 (11:00)  Fermentation 59:00 F3: OD (102.0) Contamination? (N) F3:Induction 20:00 F2: OD (130.0) Contamination? (N) F2:Induction 18:00 <Isis>               2/26/2009 (13:00)  Fermentation 61:00 F3: OD (103.0) Contamination? (N) F3:Induction 22:00 Harvest 575 ml                             F2: OD (126.7) Contamination? (N) F3:Induction 20:00 <Isis>               2/26/2009 (15:00)  Fermentation 63:00 F3: OD (-) Contamination? (N) F3:Induction 24:00 F2: OD (-) Contamination? (N) F2:Induction 22:00 Harvest 500 ml   <Isis>                2/26/2009 (17:00)  Fermentation 65:00 F3: OD (108.5) Contamination? (N) F3:Induction 26:00 F2: OD (135.2) Contamination? (N) F2:Induction 24:00

Day 3 (Feb 27 Fri)
<Isis>               2/27/2009 (11:00) Fermentation 83:00 F3: OD (114.1) Contamination? (N) F3:Induction 44:00 F2: OD (124.8) Contamination? (N) F2:Induction 42:00 <Isis>               2/27/2009 (15:00) Fermentation 87:00 F3: Harvest F3:Induction 48:00 F2: Harvest F2:Induction 46:00