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Jan Zitzmann, Tobias Weidner, Gerrit Eichner, Denise Salzig, Peter Czermak
The production of recombinant proteins in bioreactors requires real-time process monitoring and control to increase process efficiency and to meet the requirements for a comprehensive audit trail. The combination of optical near-infrared turbidity sensors and dielectric spectroscopy provides diverse system information because different measurement principles are exploited. We used this combination of techniques to monitor and control the growth and protein production of stably transformed Drosophila melanogaster S2 cells expressing antimicrobial proteins...
March 18, 2018: Sensors
Wei Du, Joeri A Jongbloets, Coco van Boxtel, Hugo Pineda Hernández, David Lips, Brett G Oliver, Klaas J Hellingwerf, Filipe Branco Dos Santos
Background: Microbial bioengineering has the potential to become a key contributor to the future development of human society by providing sustainable, novel, and cost-effective production pipelines. However, the sustained productivity of genetically engineered strains is often a challenge, as spontaneous non-producing mutants tend to grow faster and take over the population. Novel strategies to prevent this issue of strain instability are urgently needed. Results: In this study, we propose a novel strategy applicable to all microbial production systems for which a genome-scale metabolic model is available that aligns the production of native metabolites to the formation of biomass...
2018: Biotechnology for Biofuels
Gabriele Gramelsberger
Continuous culture techniques were developed in the early twentieth century to replace cumbersome studies of cell growth in batch cultures. In contrast to batch cultures, they constituted an open concept, as cells are forced to proliferate by adding new medium while cell suspension is constantly removed. During the 1940s and 1950s new devices have been designed-called "automatic syringe mechanism," "turbidostat," "chemostat," "bactogen," and "microbial auxanometer"-which allowed increasingly accurate quantitative measurements of bacterial growth...
January 30, 2018: History and Philosophy of the Life Sciences
Helmut Maske, Ramón Cajal-Medrano, Josué Villegas-Mendoza
Bacteria are the principal consumers of dissolved organic carbon (DOC) in the ocean and predation of bacteria makes organic carbon available to higher trophic levels. The efficiency with which bacteria convert the consumed carbon ( C ) into biomass (i.e., carbon growth efficiency, Y ) determines their ecological as well as biogeochemical role in marine ecosystems. Yet, it is still unclear how changes in temperature will affect Y and, hence, the transfer of consumed C to higher trophic levels. Here, we experimentally investigated the effect of temperature on metabolic functions of coastal microbial communities inoculated in both nutrient-limited chemostats and nutrient-unlimited turbidostats...
2017: Frontiers in Microbiology
Lars Fredrik Skau, Tom Andersen, Jan-Erik Thrane, Dag Olav Hessen
Temperature and nutrients are key factors affecting the growth, cell size, and physiology of marine phytoplankton. In the ocean, temperature and nutrient availability often co-vary because temperature drives vertical stratification, which further controls nutrient upwelling. This makes it difficult to disentangle the effects of temperature and nutrients on phytoplankton purely from observational studies. In this study, we carried out a factorial experiment crossing two temperatures (13°and 19°C) with two growth regimes (P-limited, semi-continuous batch cultures ["-P"] and nutrient replete batch cultures in turbidostat mode ["+P"]) for three species of common marine haptophytes (Emiliania huxleyi, Chrysochromulina rotalis and Prymnesium polylepis) to address the effects of temperature and nutrient limitation on elemental content and stoichiometry (C:N:P), total RNA, cell size, and growth rate...
2017: PeerJ
Stefan A Hoffmann, Christian Wohltat, Kristian M Müller, Katja M Arndt
For various experimental applications, microbial cultures at defined, constant densities are highly advantageous over simple batch cultures. Due to high costs, however, devices for continuous culture at freely defined densities still experience limited use. We have developed a small-scale turbidostat for research purposes, which is manufactured from inexpensive components and 3D printed parts. A high degree of spatial system integration and a graphical user interface provide user-friendly operability. The used optical density feedback control allows for constant continuous culture at a wide range of densities and offers to vary culture volume and dilution rates without additional parametrization...
2017: PloS One
Lenneke de Winter, Iago Teles Dominguez Cabanelas, Dirk E Martens, René H Wijffels, Maria J Barbosa
BACKGROUND: Day/night cycles regulate the circadian clock of organisms to program daily activities. Many species of microalgae have a synchronized cell division when grown under a day/night cycle, and synchronization might influence biomass yield and composition. Therefore, the aim of this study was to study the influence of day/night cycle on biomass yield and composition of the green microalgae Neochloris oleoabundans. Hence, we compared continuous turbidostat cultures grown under continuous light with cultures grown under simulated day/night cycles...
2017: Biotechnology for Biofuels
Christina Schreiber, Dominik Behrendt, Gregor Huber, Christian Pfaff, Janka Widzgowski, Bärbel Ackermann, Andreas Müller, Vilém Zachleder, Šárka Moudříková, Peter Mojzeš, Ulrich Schurr, Johan Grobbelaar, Ladislav Nedbal
Growth of Chlorella vulgaris was characterized as a function of irradiance in a laboratory turbidostat (1L) and compared to batch growth in sunlit modules (5-25L) of the commercial NOVAgreen photobioreactor. The effects of variable sunlight and culture density were deconvoluted by a mathematical model. The analysis showed that algal growth was light-limited due to shading by external construction elements and due to light attenuation within the algal bags. The model was also used to predict maximum biomass productivity...
June 2017: Bioresource Technology
Patrick J McGinn, Scott P MacQuarrie, Jerome Choi, Boris Tartakovsky
In this study, production of the microalga Scenedesmus AMDD in a 300 L continuous flow photobioreactor was maximized using an online flow (dilution rate) control algorithm. To enable online control, biomass concentration was estimated in real time by measuring chlorophyll-related culture fluorescence. A simple microalgae growth model was developed and used to solve the optimization problem aimed at maximizing the photobioreactor productivity. When optimally controlled, Scenedesmus AMDD culture demonstrated an average volumetric biomass productivity of 0...
January 2017: Bioprocess and Biosystems Engineering
Hugh Douglas Goold, Stéphan Cuiné, Bertrand Légeret, Yuanxue Liang, Sabine Brugière, Pascaline Auroy, Hélène Javot, Marianne Tardif, Brian Jones, Fred Beisson, Gilles Peltier, Yonghua Li-Beisson
Enriching algal biomass in energy density is an important goal in algal biotechnology. Nitrogen (N) starvation is considered the most potent trigger of oil accumulation in microalgae and has been thoroughly investigated. However, N starvation causes the slow down and eventually the arrest of biomass growth. In this study, we show that exposing a Chlamydomonas reinhardtii culture to saturating light (SL) under a nonlimiting CO2 concentration in turbidostatic photobioreactors induces a sustained accumulation of lipid droplets (LDs) without compromising growth, which results in much higher oil productivity than N starvation...
August 2016: Plant Physiology
Zinaida Eltsova, Maxim Bolshakov, Anatoly Tsygankov
The composition of photosynthetic apparatus of Rhodobacter sphaeroides wild strain 2.4.1 and its LHII-deficient mutant DBCΩ was compared. The absence of LHII in the mutant was confirmed by comparison of chromatophores spectra and by the absence of electrophoretic band corresponding to LHII complex. Continuous turbidostat cultures of wild strain and its LHII-deficient mutant were compared in response to different light intensities. Cultures were grown using lactate, mixture of lactate and acetate or succinate as carbon source...
December 2016: Photosynthesis Research
Dominick Matteau, Vincent Baby, Stéphane Pelletier, Sébastien Rodrigue
BACKGROUND: Continuous culture devices can be used for various purposes such as establishing reproducible growth conditions or maintaining cell populations under a constant environment for long periods. However, commercially available instruments are expensive, were not designed to handle small volumes in the milliliter range, and can lack the flexibility required for the diverse experimental needs found in several laboratories. METHODOLOGY/PRINCIPAL FINDINGS: We developed a versatile continuous culture system and provide detailed instructions as well as a graphical user interface software for potential users to assemble and operate their own instrument...
2015: PloS One
Pascal van Alphen, Klaas J Hellingwerf
The cyanobacterial circadian clock has been well-studied and shown to be both robust and a dominant factor in the control of gene expression in Synechococcus elongatus PCC7942. In Synechocystis sp. PCC6803, the circadian clock is assumed to function similarly, yet appears to control transcription to a far lesser extent and its circadian rhythm was reported to not be sustained, or at least rapidly damped, under continuous illumination. One of the feedback loops that governs the clock in S. elongatus in addition to the core oscillator, i...
2015: PloS One
Victoria H Work, Matthew R Melnicki, Eric A Hill, Fiona K Davies, Leo A Kucek, Alexander S Beliaev, Matthew C Posewitz
The cyanobacterium Synechococcus sp. Pasteur culture collection 7002 was genetically engineered to synthesize biofuel-compatible medium-chain fatty acids (FAs) during photoautotrophic growth. Expression of a heterologous lauroyl-acyl carrier protein (C12:0-ACP) thioesterase with concurrent deletion of the endogenous putative acyl-ACP synthetase led to secretion of transesterifiable C12:0 FA in CO2-supplemented batch cultures. When grown at steady state over a range of light intensities in a light-emitting diode turbidostat photobioreactor, the C12-secreting mutant exhibited a modest reduction in growth rate and increased O2 evolution relative to the wild-type (WT)...
2015: Frontiers in Bioengineering and Biotechnology
Ali Mohagheghi, Jeffrey G Linger, Shihui Yang, Holly Smith, Nancy Dowe, Min Zhang, Philip T Pienkos
BACKGROUND: Complete conversion of the major sugars of biomass including both the C5 and C6 sugars is critical for biofuel production processes. Several inhibitory compounds like acetate, hydroxymethylfurfural (HMF), and furfural are produced from the biomass pretreatment process leading to 'hydrolysate toxicity,' a major problem for microorganisms to achieve complete sugar utilization. Therefore, development of more robust microorganisms to utilize the sugars released from biomass under toxic environment is critical...
2015: Biotechnology for Biofuels
Hans C Bernstein, Allan Konopka, Matthew R Melnicki, Eric A Hill, Leo A Kucek, Shuyi Zhang, Gaozhong Shen, Donald A Bryant, Alexander S Beliaev
Synechococcus sp. PCC 7002 was grown to steady state in optically thin turbidostat cultures under conditions for which light quantity and quality was systematically varied by modulating the output of narrow-band LEDs. Cells were provided photons absorbed primarily by chlorophyll (680 nm) or phycocyanin (630 nm) as the organism was subjected to four distinct mono- and dichromatic regimes. During cultivation with dichromatic light, growth rates were generally proportional to the total incident irradiance at values <275 μmol photons m(-2) · s(-1) and were not affected by the ratio of 630:680 nm wavelengths...
2014: Frontiers in Microbiology
David Gresham, Maitreya J Dunham
Studying evolution in the laboratory provides a means of understanding the processes, dynamics and outcomes of adaptive evolution in precisely controlled and readily replicated conditions. The advantages of experimental evolution are maximized when the selection is well defined, which enables linking genotype, phenotype and fitness. One means of maintaining a defined selection is continuous culturing: chemostats enable the study of adaptive evolution in constant nutrient-limited environments, whereas cells in turbidostats evolve in constant nutrient abundance...
December 2014: Genomics
Chris N Takahashi, Aaron W Miller, Felix Ekness, Maitreya J Dunham, Eric Klavins
Engineered biological circuits are often disturbed by a variety of environmental factors. In batch culture, where the majority of synthetic circuit characterization occurs, environmental conditions vary as the culture matures. Turbidostats are powerful characterization tools that provide static culture environments; however, they are often expensive, especially when purchased in custom configurations, and are difficult to design and construct in a lab. Here, we present a low cost, open source multiplexed turbidostat that can be manufactured and used with minimal experience in electrical or software engineering...
January 16, 2015: ACS Synthetic Biology
Kieu-Van Dang, Julie Plet, Dimitri Tolleter, Martina Jokel, Stéphan Cuiné, Patrick Carrier, Pascaline Auroy, Pierre Richaud, Xenie Johnson, Jean Alric, Yagut Allahverdiyeva, Gilles Peltier
During oxygenic photosynthesis, metabolic reactions of CO2 fixation require more ATP than is supplied by the linear electron flow operating from photosystem II to photosystem I (PSI). Different mechanisms, such as cyclic electron flow (CEF) around PSI, have been proposed to participate in reequilibrating the ATP/NADPH balance. To determine the contribution of CEF to microalgal biomass productivity, here, we studied photosynthesis and growth performances of a knockout Chlamydomonas reinhardtii mutant (pgrl1) deficient in PROTON GRADIENT REGULATION LIKE1 (PGRL1)-mediated CEF...
July 2014: Plant Cell
K L Larsen, R P Cox, M Miller
Green photosynthetic bacteria contain a mixture of stereoisomers and homologs of their major light harvesting pigment, bacteriochlorophyll (BChl)c. We have determined the distribution of photosynthetic pigments in the green filamentous bacteriumChloroflexus aurantiacus grown in turbidostat culture under light-limited conditions at 5 different illumination intensities. Pigments were extracted from isolated cells, analyzed by HPLC, and the homologs of BChlc identified by their mass spectra. The ratio between BChlc, BChla and carotenoid remained constant at low illumination intensities; at higher intensities BChla and carotenoid increased in parallel compared to BChlc...
July 1994: Photosynthesis Research
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