TY  - JOUR
A1  - Grunzel, Petra
A1  - Pilarek, Maciej
A1  - Steinbrueck, Doerte
A1  - Neubauer, Antje
A1  - Brand, Eva
A1  - Kumke, Michael Uwe
A1  - Neubauer, Peter
A1  - Krause, Mirja
T1  - Mini-scale cultivation method enables expeditious plasmid production in Escherichia coli
JF  - Biotechnology journal : systems & synthetic biology, nanobiotech, medicine
N2  - The standard procedure in the lab for plasmid isolation usually involves a 2-mL, 16 h over-night cultivation in 15-mL bioreaction tubes in LB medium. This is time consuming, and not suitable for high-throughput applications. This study shows that it is possible to produce plasmid DNA (pDNA) in a 1.5-mL microcentrifuge tube with only 100 L cultivation volume in less than 7 h with a simple protocol. Compared with the standard LB cultivation for pDNA production reaching a final pDNA concentration range of 1.5-4 mu g mL(-1), a 6- to 10-fold increase in plasmid concentration (from 10 up to 25 mu g mL(-1) cultivation volume) is achieved using an optimized medium with an internal substrate delivery system (EnBase (R)). Different strains, plasmids, and the applicability of different inoculation tools (i.e. different starting ODs) were compared, demonstrating the robustness of the system. Additionally, dissolved oxygen was monitored in real time online, indicating that under optimized conditions oxygen limitation can be avoided. We developed a simple protocol with a significantly decreased procedure time, enabling simultaneous handling of more samples, while a consistent quality and a higher final pDNA concentration are ensured.
KW  - Escherichia coli
KW  - High-cell-density culture
KW  - Miniaturized cultivations
KW  - Optical oxygen sensor
KW  - Plasmid DNA production
Y1  - 2014
U6  - https://doi.org/10.1002/biot.201300177
SN  - 1860-6768
SN  - 1860-7314
VL  - 9
IS  - 1
SP  - 128
EP  - 136
PB  - Wiley-VCH
CY  - Weinheim
ER  -