Microscale and miniscale fermentation and screening

Lattermann, Clemens; Büchs, Jochen

doi:10.1016/j.copbio.2014.12.005

Cited by 70 publications

(47 citation statements)

References 41 publications

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“…Typically, microtiter plate readers are used to determine the optimal conditions for microbial growth and lipid production because they enable many strains and culture conditions to be screened simultaneously [20–22, 24, 47]. To this, the BioLector adds the ability to perform fermentation in conditions close to those of bench-top bioreactors, and, in addition, enables online fluorescence readings [21–23, 46].…”

Section: Resultsmentioning

confidence: 99%

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production

et al. 2016

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BackgroundBecause the model yeast Yarrowia lipolytica can synthesize and store lipids in quantities up to 20 % of its dry weight, it is a promising microorganism for oil production at an industrial scale. Typically, optimization of the lipid production process is performed in the laboratory and later scaled up for industrial production. However, the scale-up process can be complicated by genetic modifications that are optimized for one set of growing conditions can confer a less-than-optimal phenotype in a different environment. To address this issue, small cultivation systems have been developed that mimic the conditions in benchtop bioreactors. In this work, we used one such microbioreactor system, the BioLector, to develop high-throughput fermentation procedures that optimize growth and lipid accumulation in Y. lipolytica. Using this system, we were able to monitor lipid and biomass production in real time throughout the culture duration.ResultsThe BioLector can monitor the growth of Y. lipolytica in real time by evaluating scattered light; this produced accurate measurements until cultures reached an equivalent of OD600nm = 115 and a cell dry weight of 100 g L−1. In addition, a lipid-specific fluorescent probe was applied which reliably monitored lipid production up to a concentration of 12 g L−1. Through screening various growing conditions, we determined that a carbon/nitrogen ratio of 35 was the most efficient for lipid production. Further screening showed that ammonium chloride and glycerol were the most valuable nitrogen and carbon sources, respectively, for growth and lipid production. Moreover, a carbon concentration above 1 M appeared to impair growth and lipid accumulation. Finally, we used these optimized conditions to screen engineered strains of Y. lipolytica with high lipid-accumulation capability. The growth and lipid content of the strains cultivated in the BioLector were compared to those grown in benchtop bioreactors.ConclusionTo our knowledge, this is the first time that the BioLector has been used to track lipid production in real time and to monitor the growth of Y. lipolytica. The present study also showed the efficacy of the BioLector in screening growing conditions and engineered strains prior to scale-up. The method described here could be applied to other oleaginous microorganisms.

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Section: Resultsmentioning

confidence: 99%

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production

et al. 2016

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“…The most important difference between small-scale screening and bioreactor cultivation is the ability to control the specific growth rate at values below its maximum. The gap between screening in batch culture mode (in shake flasks or deep-well plates) and the subsequently performed, labour-intensive, bioreactor culture in fedbatch mode is currently addressed by small-scale bioreactors and new feeding techniques (Barnard et al, 2010, Lattermann and Buchs, 2014, Wilming et al, 2014. Downscaled techniques of substrate addition include pump-controlled feeding (Barnard et al, 2010), release of glucose from a polymer (Hemmerich et al, 2014) or by an enzyme (PanulaPerala et al, 2008), and microfluidic systems (Grunberger et al, 2014).…”

Section: Overcoming Drawbacks Of Current Screening Conceptsmentioning

confidence: 98%

Cultivation strategies to enhance productivity of Pichia pastoris: A review

Looser

Brühlmann

Bumbak

et al. 2015

Biotechnology Advances

283

268

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Pichia pastoris, a methylotrophic yeast, is an established system for the production of heterologous proteins, particularly biopharmaceuticals and industrial enzymes. To maximise and optimise the production of recombinant products, recent molecular research has focused on numerous issues including the design of expression vectors, optimisation of gene copy number, co-expression of secretory proteins such as chaperones, engineering of glycosylation and secretory pathways, etc. However, the physiological effects of different cultivation strategies are often difficult to separate from the molecular effects of the gene construct (e.g., cellular stress through over-expression or incorrect post-translational processing). Hence, overall system optimisation is difficult, even though it is urgently required in order to describe and understand the behaviour of new molecular constructs. This review focuses on particular aspects of recombinant protein production related to variations in biomass growth and their implications for strain design and screening, as well as on the concept of rational comparisons between cultivation systems for the development of specific production processes in bioreactors. The relationship between specific formation rates of secreted recombinant proteins, qp, and specific growth rates, μ, has been analysed in a conceptual attempt to compare different systems, particularly those based on AOX1/methanol and GAP/glucose, and this has now evolved into a pivotal concept for bioprocess engineering of P. pastoris.

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“…Hegab et al (2013) gave an overview of MBR fabrication techniques as well as their operation and control. Lattermann and Büchs (2015) reviewed miniaturized bioreactors and MBR development, highlighting the mass transfer and power input characterization, optical monitoring, and automation of fed-batch screening systems. Lattermann and Büchs (2015) reviewed miniaturized bioreactors and MBR development, highlighting the mass transfer and power input characterization, optical monitoring, and automation of fed-batch screening systems.…”

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confidence: 99%

A fully online sensor‐equipped, disposable multiphase microbioreactor as a screening platform for biotechnological applications

Maldonado

Panjan

Sun

et al. 2018

Biotech & Bioengineering

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A new disposable, multiphase, microbioreactor (MBR; with a working volume of 550 μl) equipped with online sensors is presented for biotechnological screening research purposes owing to its high-throughput potential. Its design and fabrication, online sensor integration, and operation are described. During aerobic cultivation, sufficient oxygen supply is the most important factor that influences growth and product formation. The MBR is a microbubble column bioreactor (μBC), and the oxygen supply was realized by active pneumatic bubble aeration, ensuring sufficient volumetric liquid-phase mass transfer (k a) and proper homogenization of the cultivation broth. The μBC was equipped with miniaturized sensors for the pH, dissolved oxygen, optical density and glucose concentration that allowed real-time online monitoring of these process variables during cultivation. The challenge addressed here was the integration of sensors in the limited available space. The MBR was shown to be a suitable screening platform for the cultivation of biological systems. Batch cultivations of Saccharomyces cerevisiae were performed to observe the variation in the process variables over time and to show the robustness and operability of all the online sensors in the MBR.

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Microscale and miniscale fermentation and screening

Cited by 70 publications

References 41 publications

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production

Cultivation strategies to enhance productivity of Pichia pastoris: A review

A fully online sensor‐equipped, disposable multiphase microbioreactor as a screening platform for biotechnological applications

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