133 research outputs found
Establishing a high throughput method for medium optimization â a case study using Streptomyces lividans as host for production of heterologous protein
Synthetic Promoter Library for modulation of actinorhodin production in Streptomyces coelicolorA3(2)
Commissioning and equipment assessment of a semi-industrial bioreactor
Tasks regarding the commission and equipment assessment on the fermenter situated in PILOT PLANT Research Center were carried out. The bioreactor consists in a pilotscale fermenter with a capacity of 150L. Piping and Instrumentation Diagrams were elaborated, in which a posterior Sterilization Standard Operating Procedure was designed. In such SOP, Steam In Place method was implemented. The sterilization was designed as a batch process, in which the reaction medium is sterilized simultaneously to the fermentation vessel. This way, the use of additional equipment was avoided and the risk of contamination between sterilization and fermentation start was minimized. Optimal saturated vapor temperature was taken as 121ÂşC. The required holding time for equipment sterilization (spare parts) was determined to be 10 minutes, whereas a holding time of 20 minutes was considered for medium sterilization. Other key factors taken into account for the design of the procedure were condensate removal, air evacuation and post-sterilization integrity, The SOP is still in need of validation. A sealing test revealed the inoperability of the safety device (burst disk). This device was then replaced and tested with success. Gas-liquid mass transfer capacity of the fermenter was assessed, applying the hydrogen peroxide method. A KLa of 25Âą3.3 h-1was determined when the stirrer run at 450 rpm. However, due to supersaturation effect, this value could be an overestimation of the actual value. Comparison with other typical KLa values for pilot-scale bioreactors led to the conclusion that the aeration capacity of the fermenter is low and might be insufficient for some biochemical processes. A change in the impeller type is proposed in order to address this problem
Experimental and in-silico investigation of population heterogeneity in continuous Sachharomyces cerevisiae scale-down fermentation in a novel two-compartment setup.
Robust, small-scale cultivation platform for Streptomyces coelicolor
<p>Abstract</p> <p>Background</p> <p>For fermentation process and strain improvement, where one wants to screen a large number of conditions and strains, robust and scalable high-throughput cultivation systems are crucial. Often, the time lag between bench-scale cultivations to production largely depends on approximate estimation of scalable physiological traits. Microtiter plate (MTP) based screening platforms have lately become an attractive alternative to shake flasks mainly because of the ease of automation. However, there are very few reports on applications for filamentous organisms; as well as efforts towards systematic validation of physiological behavior compared to larger scale are sparse. Moreover, available small-scale screening approaches are typically constrained by evaluating only an end point snapshot of phenotypes.</p> <p>Results</p> <p>To address these issues, we devised a robust, small-scale cultivation platform in the form of MTPs (24-square deepwell) for the filamentous bacterium <it>Streptomyces coelicolor </it>and compared its performance to that of shake flasks and bench-scale reactors. We observed that re-designing of medium and inoculum preparation recipes resulted in improved reproducibility. Process turnaround time was significantly reduced due to the reduction in number of unit operations from inoculum to cultivation. The incorporation of glass beads (ø 3 mm) in MTPs not only improved the process performance in terms of improved oxygen transfer improving secondary metabolite production, but also helped to transform morphology from pellet to disperse, resulting in enhanced reproducibility. Addition of MOPS into the medium resulted in pH maintenance above 6.50, a crucial parameter towards reproducibility. Moreover, the entire trajectory of the process was analyzed for compatibility with bench-scale reactors. The MTP cultivations were found to behave similar to bench-scale in terms of growth rate, productivity and substrate uptake rate and so was the onset of antibiotic synthesis. Shake flask cultivations however, showed discrepancy with respect to morphology and had considerably reduced volumetric production rates of antibiotics.</p> <p>Conclusion</p> <p>We observed good agreement of the physiological data obtained in the developed MTP platform with bench-scale. Hence, the described MTP-based screening platform has a high potential for investigation of secondary metabolite biosynthesis in <it>Streptomycetes </it>and other filamentous bacteria and the use may significantly reduce the workload and costs.</p
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