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Publication
Featured researches published by Bianca Elisabeth Maria Gielesen.
Archive | 2013
Bianca Elisabeth Maria Gielesen; Marco van den Berg
The polyethylene glycol (PEG) protoplast method for transformation of filamentous fungi is developed for use in microtiter plates, enabling one to perform a multitude of transformations in parallel.
Metabolic Engineering | 2018
Eric M. Young; Zheng Zhao; Bianca Elisabeth Maria Gielesen; Liang Wu; D. Benjamin Gordon; Johannes Andries Roubos; Christopher A. Voigt
Metabolic engineering requires multiple rounds of strain construction to evaluate alternative pathways and enzyme concentrations. Optimizing multigene pathways stepwise or by randomly selecting enzymes and expression levels is inefficient. Here, we apply methods from design of experiments (DOE) to guide the construction of strain libraries from which the maximum information can be extracted without sampling every possible combination. We use Saccharomyces cerevisiae as a host for a novel six-gene pathway to itaconic acid, selected by comparing alternative shunt pathways that bypass the mitochondrial TCA cycle. The pathway is distinctive for the use of acetylating acetaldehyde dehydrogenase to increase cytosolic acetyl-CoA pools, a bacterial enzyme to synthesize citrate in the cytosol, and an itaconic acid exporter. Precise control over the expression of each gene is enabled by a set of promoter-terminator pairs that span a 174-fold range. Two large combinatorial libraries (160 variants, 2.4 Mb and 32 variants, 0.6 Mb) are designed where the expression levels are selected by statistical methods (I-optimal response surface methodology, full factorial, or Plackett-Burman) with the intent of extracting different types of guiding information after the screen. This is applied to the design of a third library (24 variants, 0.5 Mb) intended to alleviate a bottleneck in cis-aconitate decarboxylase (CAD) expression. The top strain produces 815 mg/l itaconic acid, a 4-fold improvement over the initial strain achieved by iteratively balancing pathway expression. Including a methylated product in the total, the strain produces 1.3 g/l combined itaconic acids. Further, a regression analysis of the libraries reveals the optimal expression level of CAD as well as pairwise interdependencies between genes that result in increased titer and purity of itaconic acid. This work demonstrates adapting algorithmic design strategies to guide automated yeast strain construction and learn information after each iteration.
Archive | 2009
Paul Klaassen; Van Der Jan Metske Laan; Bianca Elisabeth Maria Gielesen; Van Gijsberdina Pieternella Suylekom
Archive | 2010
Paul Klaassen; Van Gijsberdina Pieternella Suylekom; Bianca Elisabeth Maria Gielesen; Nicolette Jasmijn Broers; Beate Wiedemann; De Wilhelmus Theodorus Antonius Maria Laat
Archive | 2011
Paul Klaassen; Bianca Elisabeth Maria Gielesen; Van Gijsberdina Pieternella Suylekom; Wilbert Herman Marie Heijne
Archive | 2011
Paul Klaassen; Bianca Elisabeth Maria Gielesen; Wilbert Herman Marie Heijne; Van Gijsberdina Pieternella Suylekom
Archive | 2012
Paul Klaassen; Bianca Elisabeth Maria Gielesen; Gijsberdina Pieternella Van Suylekom; Panagiotis Sarantinopoulos; Wilbert Herman Marie Heijne; Aldo Greeve
Archive | 2007
Marco van den Berg; Bianca Elisabeth Maria Gielesen; Adriana Marina Riemens
Archive | 2016
Bernard Meijrink; René Verwaal; Bianca Elisabeth Maria Gielesen; Johannes Andries Roubos
Archive | 2016
Bernard Meijrink; René Verwaal; Bianca Elisabeth Maria Gielesen; Johannes Andries Roubos; Yvonne Nygård; Roelof Arij Lans Bovenberg; Arnold J. M. Driessen; Carsten Pohl