User:Fermenter User/Notebook/BROM A17

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BROM A17


Day -2 (Nov 14 Sun):

      <Dennis>           11/8/2010 ()
                               Thaw glycerol stock and grow in 5-ml MD media (prepared in summertime)
                               Incubator in Fermentation Suite used
      <Dennis>         11/8/2010 (before go home)
                              Preparing 5-L of Basal Medium (Defined media)
                              Ortho-phosphoric acid is borrowed from Molday lab
                              PTM1 Salt is from BROM-A16
Freshly filtered PTM salt <-- SAME as BROM A16
Freshly filtered PTM salt <-- SAME as BROM A16
Defined Media for 5-L
Defined Media for 5-L

Day -1 (Nov 15 Mon): MEDIA PREPARATION DAY

      <Sung-Hye>         11/15/2010 (14:00)    Assemble bioreactors:   
                              1. Calibrate pH sensors                                         
                              2. 5.0-L of Basal Salts:  
                              3. 1-ml Antifoam 
<Sung-Hye> 11/15/2010 (14:00) Prepare bottles 1) no Acid need 2) Base, 500-ml (tubing connected) x2 3) MeOH, 1-L (tubing connected) x2 4) Additives/Inoculum, 1 x 500-ml (tubing connected) x2 5) Water, 2-L

<Sung-Hye> 11/15/2010 (14:30) Autoclave <Sung-Hye> 11/15/2010 (16:00) Autoclave completed! <--Door stuck! <Dennis> 11/15/2010 (11:50 - 12:00) Transfer cells to fresh 55-ml BMGY media Incubator in Fermentation Suite used

Day 0 (Nov 16 Tue): INOCULATION DAY

      <Sung-Hye>         11/16/2010 (9:45)   Set temperature to 28.5°C 
      <Sung-Hye>         11/16/2010 (9:45)   Purge Air 1L/min at 900 rpm 
      <Sung-Hye>         11/16/2010 (9:50)   Adjust pH to 5.0 with 140 ml NH4OH 
      <Sung-Hye>         11/16/2010 (9:50)      Start pH control 
                                                  F1: No Acid Base, 1M (400 ml)
      <Sung-Hye>         11/16/2010 (9:51)      Start Fermenter #1 software 
                         11/16/2010 (10:00)      Calibrate dO2 probes 
                         11/16/2010 (10:00)      Strt dO2 control 
<Dennis> 11/16/2010 (12:30) Check contamination of starter culture
<Dennis&Sung-Hye> 11/16/2010 (13:00) Fermentation 0:00 Flask 1 (from frozen stock) OD: 15 Inoculate 45 ml/bioreactor <Sung-Hye> 11/16/2010 (16:30) Add 22 ml of PTM1 Salt/bioreactor

Day 1 (Nov 17 Wed):

      <Dennis>           11/17/2010 (12:00)      Fermentation 23:00  
                            F1 Base(400 ml) OD(1.9) Contamination(N) Pressure(N) Change Filter(N)

Day 2 (Nov 18 Thu)

      <Dennis>           11/18/2010 (8:00)      Fermentation 44:00  
                            F1 Base(400 ml) OD(8.1) Contamination(N) Pressure(N) Change Filter(N)
      <Dennis>           11/18/2010 (15:00)     Fermentation 51:00  
                            F1 Base(400 ml) OD(10.75) Contamination(N) Pressure(N) Change Filter(N)

Day 3 (Nov 19 Fri): MeOH Induction Day 1

      <Sung-Hye>         11/19/2010 (09:00)     Fermentation 69:00  
                            Base (0 ml)
                            pH <5.0 (Adjust pH to 5.0 using conc. NH4OH
                            Refill Base (+400 ml with 2M)
                            Remove 500-ml media
                            Change filter                       
                             
      <Dennis>           11/19/2010 (12:00)      Fermentation 71:00  
                            F1: Base(?? ml) OD(51.25) Contamination(N) Pressure(N) Change Filter(N)
      <Sung-Hye>         11/19/2010 (13:00)      Fermentation 73:00  MeOH induction 00:00 
                            Calibrate MeOH sensor  6287 mV = 0.5%
                            Start MeOH induction

Day 4 (Nov 20 Sat): MeOH Induction Day 2

      <Dennis>           11/20/2010 (14:00)      Fermentation 96:00  MeOH induction 24:00  
                            F1: Base(350 ml) MeOH(450 ml) OD(45.3) Contamination(N) Pressure(N) Change Filter(N)
                                Activity assay (9.24 units/100ul), (2.16 mg/5-L)

Day 5 (Nov 21 Sun): MeOH Induction Day 3

      <Dennis>           11/21/2010 (14:00)      Fermentation 120:00  MeOH induction 48:00  
                            F1: Base(350 ml) MeOH(425 ml) OD(46.6) Contamination(N) Pressure(N) Change Filter(N)
                                Activity assay (21.49 units/100ul), (5.03 mg/5-L)

Day 6 (Nov 22 Mon): MeOH Induction Day 4

      <Dennis>           11/22/2010 (14:00)      Fermentation 144:00  MeOH induction 72:00  
                            F1: Base(340 ml) MeOH(400 ml) OD(49.25) Contamination(N) Pressure(N) Change Filter(N)
                                Activity assay (23.03 units/100ul), (5.39 mg/5-L)

Day 7 (Nov 23 Tue): MeOH Induction Day 5

      <Dennis>           11/23/2010 (14:00)      Fermentation 168:00  MeOH induction 96:00  
                            F1: Base(300 ml) MeOH(375 ml) OD(49.0) Contamination(N) Pressure(N) Change Filter(N)
                                Activity assay (21 units/100ul), (4.94 mg/5-L)
                                Harvest



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