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Development of an optimized medium, strain and high-throughput culturing methods for Methylobacterium extorquens.

Authors :
Nigel F Delaney
Maria E Kaczmarek
Lewis M Ward
Paige K Swanson
Ming-Chun Lee
Christopher J Marx
Source :
PLoS ONE, Vol 8, Iss 4, p e62957 (2013)
Publication Year :
2013
Publisher :
Public Library of Science (PLoS), 2013.

Abstract

Methylobacterium extorquens strains are the best-studied methylotrophic model system, and their metabolism of single carbon compounds has been studied for over 50 years. Here we develop a new system for high-throughput batch culture of M. extorquens in microtiter plates by jointly optimizing the properties of the organism, the growth media and the culturing system. After removing cellulose synthase genes in M. extorquens strains AM1 and PA1 to prevent biofilm formation, we found that currently available lab automation equipment, integrated and managed by open source software, makes possible reliable estimates of the exponential growth rate. Using this system, we developed an optimized growth medium for M. extorquens using response surface methodologies. We found that media that used EDTA as a metal chelator inhibited growth and led to inconsistent culture conditions. In contrast, the new medium we developed with a PIPES buffer and metals chelated by citrate allowed for fast and more consistent growth rates. This new Methylobacterium PIPES ('MP') medium was also robust to large deviations in its component ingredients which avoided batch effects from experiments that used media prepared at different times. MP medium allows for faster and more consistent growth than other media used for M. extorquens.

Subjects

Subjects :
Medicine
Science

Details

Language :
English
ISSN :
19326203
Volume :
8
Issue :
4
Database :
Directory of Open Access Journals
Journal :
PLoS ONE
Publication Type :
Academic Journal
Accession number :
edsdoj.be3a2c4c066c4c488611be5f7eaf5343
Document Type :
article
Full Text :
https://doi.org/10.1371/journal.pone.0062957