% IMPORTANT: The following is UTF-8 encoded. This means that in the presence
% of non-ASCII characters, it will not work with BibTeX 0.99 or older.
% Instead, you should use an up-to-date BibTeX implementation like “bibtex8” or
% “biber”.
@PHDTHESIS{Hoffmann:995158,
author = {Hoffmann, Kyra},
othercontributors = {Büchs, Jochen and Conrath, Uwe},
title = {{O}nline analysis of microbial γ-polyglutamic acid
production and plant defense priming in small-scale cell
culture},
school = {Rheinisch-Westfälische Technische Hochschule Aachen},
type = {Dissertation},
address = {Aachen},
publisher = {RWTH Aachen University},
reportid = {RWTH-2024-09757},
pages = {1 Online-Ressource : Illustrationen},
year = {2024},
note = {Veröffentlicht auf dem Publikationsserver der RWTH Aachen
University; Dissertation, Rheinisch-Westfälische Technische
Hochschule Aachen, 2024},
abstract = {The development of online analytical tools significantly
accelerates research of bioprocesses in small shaken
cultures. This thesis demonstrates the application of online
analytics for two processes. Firstly for the production of
poly--glutamic acid (γ-PGA) in shake flasks and secondly
for the detection of priming-active compounds with parsley
cell cultures in microtiter plates. These two diverse
research topics demonstrate the power of online analytic
tools in small culture scale. The first chapter of this
thesis is about the online screening of γ-PGA, a biopolymer
with a wide range of applications. The γ-PGA was produced
with genetically modified Bacillus subtilis 168 strains. The
formation and concomitant secretion of γ-PGA increase the
culture broth viscosity, while enzymatic depolymerization
and degradation of γ-PGA by native depolymerases decrease
the viscosity. The viscosity monitoring online system
(ViMOS) was applied for optical online measurements of broth
viscosity in shake flasks. This online monitoring enabled
the analysis of the promoter Ppst in combination with
different γ PGA depolymerase and by-product knockout
mutants in genetically modified B. subtilis 168 strains. The
different single depolymerase knockout mutants were ∆ggt,
∆pgdS, ∆cwlO, and a triple knockout mutant.
Additionally, to the Ppst promoter, the γ-PGA production
under the control of the Pxyl promoter was analyzed. An
increase in γ-PGA yield in gγ PGA/gglucose of $190\%$
could be achieved with the triple knockout mutant compared
to the Ppst reference strain. The strain with the Pxyl
promoter produced even 8.4 gγ-PGA/molcarbon. This is a
higher γ-PGA yield than achieved using the glutamic
acid-dependent γ-PGA producer Bacillus licheniformis ATCC
9945. Therefore, Bacillus subtilis Pxyl is a promising
candidate for γ-PGA production with glucose as the sole
carbon source. The second chapter of this thesis presents a
biochemical approach to detect defense priming in parsley
cell cultures in microtiter plates. Conventional crop
protection has major drawbacks, such as developing pest and
pathogen insensitivity to pesticides and low environmental
compatibility. Therefore, alternative crop protection
strategies are needed. One approach treats crops with
chemical compounds that induce a primed state of enhanced
defense. Online identification of priming-inducing chemistry
by recording breathing activity with the oxygen transfer
rate (OTR) represents a highly informative approach to spot
priming-activating chemistry. This thesis shows the online
measurement of the dissolved oxygen tension (DOT) in
microtiter plates (MTPs) and the calculation of the OTR from
the DOT to be a valid tool to draw conclusions regarding
priming-inducing activity. Furthermore, the fluorescence of
the priming-active reference compound salicylic acid (SA)
and furanocoumarins was simultaneously monitored online in
MTPs. Determining the OTR, fluorescence of the
priming-active chemical compound SA, and furanocoumarins in
parsley suspension cultures in MTPs by online measurement
allows an in-depth screening for priming compounds and a
better understanding of the priming process induced by a
given substance.},
cin = {416510 / 161620 / 161710},
ddc = {620},
cid = {$I:(DE-82)416510_20140620$ / $I:(DE-82)161620_20140620$ /
$I:(DE-82)161710_20140620$},
pnm = {SeleCa - Selectivity in Chemo- and Biocatalysis (SeleCa)
(SELECA-20170406) / Doktorandenprogramm
(PHD-PROGRAM-20170404)},
pid = {G:(DE-Juel1)SELECA-20170406 /
G:(DE-HGF)PHD-PROGRAM-20170404},
typ = {PUB:(DE-HGF)11},
doi = {10.18154/RWTH-2024-09757},
url = {https://publications.rwth-aachen.de/record/995158},
}
guest ::