Mitglied der Helmholtz-Gemeinschaft Launching an automated microtiter cultivation platform for enhanced bioprocess optimization 18th April 2013 | Forschungszentrum Jülich GmbH IBG-1: Biotechnology Institute of Bio- and Geosciences (IBG) Bioprocesses & Bioanalytics Group Leo-Brandt-Str. / 52428 Jülich / Germany www.fz-juelich.de/ibg/ibg-1 P. Rohe, O. Schweissgut, R. Freudl, W. Wiechert, M. Oldiges
26
Embed
Launching an automated microtiter cultivation platform for ... an automated microtiter cultivation platform for enhanced bioprocess optimization 18th April 2013 | F ... Optimizing
This document is posted to help you gain knowledge. Please leave a comment to let me know what you think about it! Share it to your friends and learn new things together.
Transcript
Mitg
lied
der H
elm
holtz
-Gem
eins
chaf
t
Launching an automated microtiter cultivation platform for enhanced bioprocess optimization
18th April 2013 | Forschungszentrum Jülich GmbH IBG-1: Biotechnology Institute of Bio- and Geosciences (IBG) Bioprocesses & Bioanalytics Group Leo-Brandt-Str. / 52428 Jülich / Germany www.fz-juelich.de/ibg/ibg-1
P. Rohe, O. Schweissgut, R. Freudl, W. Wiechert, M. Oldiges
Folie 2
Outline
Introduction: What is so complicated about heterologous protein expression and why do we need higher troughput ? Results:
• JuBOS: Juelich Bioprocess Optimization System – A smart platform for small scale cultivation • Bioprocess optimization of C. glutamicum as expression system
Introduction: What is so complicated about heterologous protein expression and why do we need higher troughput ? Results:
• JuBOS: Juelich Bioprocess Optimization System – A smart platform for small scale cultivation • Bioprocess optimization of C. glutamicum as expression system
• Installed in laminar flow cabinet • Automated media preparation • Pipetting (sampling/dosing/harvest)
• Programmable pipetting events triggered by (CDW, pO2, Time, pH) • Glucose fed-batch medium
Slide 7
Outline
Introduction: What is so complicated about heterologous protein expression and why do we need higher troughput ? Results:
• JuBOS: Juelich Bioprocess Optimization System – A smart platform for small scale cultivation • Bioprocess optimization of C. glutamicum as expression system
Classical and MTP-based glucose feedrate optimization
Premliminary tests
Juelich Bioprocess Optimization System (JuBOS)
Slide 22
Start Feed
0
10
20
30
40
50
60
70
0 20 40 60
EA [k
U/L
]
CDW [g.L-1]
0
1
2
3
4
0 5 10 15
EAsp
ec [k
U/g
] Feed [gGlc
.L-1.h-1]
Start Feed
benchmark (batch)
DO
[%]
NprE-Cutinase
Classical feedrate optimization in 1 L bioreactor
Glucose feed rate is a critical design parameter for fed-batch Performance increase (3x) with optimal fed-batch setup
Different signal peptides showed different optimal fed-batch feedrates !
Slide 23
Strain screening
media optimization
induction optimization
feedrate screening
feed optimi- zation
Summary: Bioprocess optimization of cutinase expression in C. glutamicum using JuBOS
Premliminary tests
scale-up
14 days 4 days 16 days 4 days
240 cultivations
96 cultivations
384 cultivations
96 cultivations
Bioprocess optimization with 816 cultivations Possible with 1 person in ~6 weeks in MTP based cultivations
Juelich Bioprocess Optimization System (JuBOS)
Slide 24
Summary: Bioprocess optimization of cutinase expression in C. glutamicum using JuBOS
Strain screening
media optimization
induction optimization
feedrate screening
Premliminary tests
• Severe cross-relationship between different optimization modules oberved
• Initial strain screening conditions must be similar to production conditions
• Arrangement of the modules have direct effect on the final optimal set of biological and bioprocess parameters (local vs. global optimum) • Finding optimal standard conditions is a very unlikely scenario for protein secretion in C. glutamicum (also in other microbial systems ?) Strong demand for target protein specific de-novo optimization approach ! • JuBOS setup provide powerful technical environment for enhanced bioprocesss optimization under conditions very similar to lab-sale bioreactor conditions Outlook: • Implementation of parallelized fed-batch option and pH control in micro plate Strong demand for design of experiments
Folie 25
Acknowledgements
Forschungszentrum Jülich (IBG-1) : Prof. Bott Dr. Britta Kleine
Dr. Frank Kensy Dipl.-Ing. Carsten Müller
Project partners:
Heinrich-Heine University Düsseldorf: Prof. K.E. Jäger Prof. Pietruzska Dr. Ulrich Krauss Dr. Britta Kleine Kathrin Scholz