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Biotechnology Journal

Sanelle van Wyk, Filipa V M Silva
Brettanomyces yeasts are increasingly being used to produce lambic style beers and craft beers with unique flavours. Currently, the industry monitors Brettanomyces bruxellensis using time consuming plate counting. B. bruxellensis is a fastidious slow growing organism, requiring 5 days of incubation at 30 °C for visible growth on agar plates. Thus, a need exists to develop a quicker, feasible method to enumerate this yeast. The aim of this study was therefore to determine the feasibility of using the 'direct' and 'indirect' impedance methods for the enumeration of B...
October 25, 2016: Biotechnology Journal
Gaia Pigna, Taniya Dhillon, Elizabeth M Dlugosz, Joshua S Yuan, Connor Gorman, Piero Morandini, Scott C Lenaghan, C Neal Stewart
Arundo donax L. is a promising biofuel feedstock in the Mediterranean region. Despite considerable interest in its genetic improvement, Arundo tissue culture and transformation remains arduous. We developed methodologies for cell- and tissue culture and genetic engineering in Arundo. A media screen was conducted, and a suspension culture was established using callus induced from stem axillary bud explants. DBAP medium, containing 9 μM 2,4-D and 4.4 μM BAP, was found to be the most effective medium among those tested for inducing cell suspension cultures, which resulted in a 5-fold increase in tissue mass over 14 days...
October 20, 2016: Biotechnology Journal
Cláudia C Miranda, Tiago G Fernandes, M Margarida Diogo, Joaquim M S Cabral
The demand of high cell numbers for applications in cellular therapies and drug screening requires the development of scalable platforms capable to generating highly pure populations of tissue-specific cells from human pluripotent stem cells. In this work, we describe the scaling-up of an aggregate-based culture system for neural induction of human induced pluripotent stem cells (hiPSCs) under chemically-defined conditions. A combination of non-enzymatic dissociation and rotary agitation was successfully used to produce homogeneous populations of hiPSC aggregates with an optimal (140 μm) and narrow distribution of diameters (coefficient of variation of 21...
October 18, 2016: Biotechnology Journal
Beate Hintersteiner, Nico Lingg, Evelyne Janzek, Oliver Mutschlechner, Hans Loibner, Alois Jungbauer
It has previously been shown for individual antibodies, that the microheterogenity pattern can have a significant impact on various key characteristics of the product. The aim of this study to get a more generalized understanding of the importance of microheterogeneity. For that purpose, the charge variant pattern of various different commercially available therapeutic mAb products was compared using Cation-Exchange Chromatography with linear pH gradient antigen affinity, Fc-receptor affinity, antibody dependent cellular cytotoxicity (ADCC) and conformational stability...
October 18, 2016: Biotechnology Journal
Jianhua Yang, Anna Joëlle Ruff, Stefanie Nicole Hamer, Feng Cheng, Ulrich Schwaneberg
Escherichia coli is a common host for recombinant protein production in which production titers are highly dependent on the employed expression system. Promoters are thereby a key element to control gene expression levels. In this study, a novel PLICable promoter toolbox was developed which enables in a single cloning step and after a screening experiment to identify out of ten IPTG-inducible promoters (T7, A3, lpp, tac, pac, Sp6, lac, npr, trc and syn) the most suitable one for high level protein production...
October 18, 2016: Biotechnology Journal
Henning Gram Hansen, Helene Faustrup Kildegaard, Gyun Min Lee, Stefan Stefan Kol
Accurate titer determination of recombinant proteins is crucial for evaluating protein production cell lines and processes. Even though enzyme-linked immunosorbent assay (ELISA) is the most widely used assay for determining protein titer, little is known about the accuracy of commercially available ELISA kits. We observed that estimations of recombinant human ø1-antitrypsin (rø1AT) titer by Coomassie-stained SDS-PAGE gels did not correspond to previously obtained titers obtained by a commercially available ELISA kit...
October 12, 2016: Biotechnology Journal
Shuhei Yamamoto, Kazunori Shimizu, Jiahui Fei, Hiroji Iwata, Mina Okochi, Hayao Nakanishi, Hiroyuki Honda
Phenotype-based analysis of circulating tumor cells (CTCs) is a promising approach to identification of new therapeutic targets and to elucidation of the biological properties. Nonetheless, ex vivo culturing of CTCs is still a technically challenging. Here, we developed a novel ex vivo culture method for CTCs using a fibroblast feeder layer and a magnetic coculture protocol. CTCs in the blood of a mouse metastasis model were labeled magnetically with magnetite nanoparticles. The labeled CTCs were isolated by a magnetic capture column and a size-selective capture filter...
October 12, 2016: Biotechnology Journal
Caroline Loy, Audrey Lainé, Diego Mantovani
Type I collagen gel is often used as a tubular scaffold because of its easy molding properties as well as its biocompatibility, low immunogenicity and ability to be remodelled by cells. However, its highly hydrated structure contributes to its weak mechanical properties and reduces its ability to be handled, which is important in tubular tissue engineering. Although cell-driven remodelling of collagen matrices is known to reinforce their mechanical properties, this process can take weeks. This study introduces a novel, simple, and rapid technique using a rotational bioreactor to expel water and densify collagen under sterile conditions to generate denser and stronger collagen gel scaffolds...
October 12, 2016: Biotechnology Journal
I-Ming Chu
No abstract text is available yet for this article.
October 12, 2016: Biotechnology Journal
Julia Hilscher, Hermann Bürstmayr, Eva Stoger
The development of gene targeting and gene editing techniques based on programmable site-directed nucleases (SDNs) has increased the precision of genome modification and made the outcomes more predictable and controllable. These approaches have achieved rapid advances in plant biotechnology, particularly the development of improved crop varieties. Here, we review the range of alterations which have already been implemented in plant genomes, and summarize the reported efficiencies of precise genome modification...
October 11, 2016: Biotechnology Journal
Lu-Hong Lan, Han Zhao, Jin-Chun Chen, Guo-Qiang Chen
Halomonas spp. have been studied as a low cost production host for producing bulk materials such as polyhydroxyalkanoates (PHA) bioplastics, since they are able to grow at high pH and high NaCl concentration under unsterile and continuous conditions without microbial contamination. In this paper, Halomonas strain TD was used as a host to produce a protein named PHA phasin or PhaP which has a potential to be developed into a bio-surfactant. Four Halomonas TD expression strains were constructed based on a strong T7-family expression system...
September 30, 2016: Biotechnology Journal
Antonino Biundo, Doris Ribitsch, Georg Steinkellner, Karl Gruber, Georg M Guebitz
An esterase from Clostridium botulinum (Cbotu_EstA) previously reported to hydrolyze the biodegradable polyester poly(butylene adipate-co-terephthalate) was redesigned to improve the hydrolysis of synthetic polyesters. Increased activity was indeed observed for del71Cbotu_EstA variant, which performed activity on the widespread polyester polyethylene terephthalate, which was not able to be attacked by the wild-type enzyme Cbotu_EstA. Analysis of the 3D structure of the enzyme showed that removing 71 residues at the N-terminus of the enzyme exposed a hydrophobic patch on the surface and improved sorption of hydrophobic polyesters concomitantly facilitating the access of the polymer to the active site...
September 30, 2016: Biotechnology Journal
Sabine Fräbel, Markus Krischke, Agata Staniek, Heribert Warzecha
Halogenation of natural compounds in planta is rare. Herein, we describe successful engineering of tryptophan 6-halogenation into the plant context by heterologous expression of the Streptomyces toxytricini Stth gene and localization of its enzymatic product in various tobacco cell compartments. When co-expressed with the flavin reductase rebF from Lechevalieria aerocolonigenes, Stth efficiently produced chlorinated tryptophan in the cytosol. Further, supplementation of KBr yielded the brominated metabolite...
September 30, 2016: Biotechnology Journal
Timothy M Pabst, Michaela Wendeler, Xiangyang Wang, Sandra Bezemer, Pim Hermans, Alan K Hunter
Interest in new and diverse classes of molecules such as recombinant toxins, enzymes, and blood factors continues to grow for use a biotherapeutics. Compared to monoclonal antibodies, these novel drugs typically lack a commercially available affinity chromatography option, which leads to greater process complexity, longer development timelines, and poor platformability. To date, for both monoclonal antibodies and novel molecules, affinity chromatography has been mostly reserved for separation of process-related impurities such as host cell proteins and DNA...
September 27, 2016: Biotechnology Journal
Carlos H Luna-Flores, Robin W Palfreyman, Jens O Krömer, Lars K Nielsen, Esteban Marcellin
Traditionally derived from fossil fuels, biological production of propionic acid has recently gained interest. Propionibacterium species produce propionic acid as their main fermentation product. Production of other organic acids reduces propionic acid yield and productivity, pointing to by-products gene-knockout strategies as a logical solution to increase yield. However, removing by-product formation has seen limited success due to our inability to genetically engineer the best producing strains (i.e. Propionibacterium acidipropionici)...
September 27, 2016: Biotechnology Journal
Tomokazu Yada, Koichi Nonaka, Masayuki Yabuta, Noriko Yoshimoto, Shuichi Yamamoto
Protein A chromatography (PAC) is commonly used as an efficient capture step in monoclonal antibody (mAb) separation processes. Usually dynamic binding capacity is used for choosing the right PAC. However, if aggregates can be efficiently removed during elution, it can make the following polishing steps easier. In this study a method for choosing the right PAC media in terms of mAb aggregate removal is proposed. Linear pH gradient elution experiments of two different mAbs on various PAC columns are carried out, where the elution behavior of aggregates as well as the monomer is measured...
September 23, 2016: Biotechnology Journal
Eva Brauchle, Katja Schenke-Layland
No abstract text is available yet for this article.
September 23, 2016: Biotechnology Journal
Ruben R G Soares, Daniel F C Silva, Pedro Fernandes, Ana M Azevedo, Virginia Chu, João P Conde, M Raquel Aires-Barros
Aqueous two-phase extraction (ATPE) is a biocompatible liquid-liquid (L-L) separation technique that has been under research for several decades towards the purification of biomolecules, ranging from small metabolites to large animal cells. More recently, with the emergence of rapid-prototyping techniques for fabrication of microfluidic structures with intricate designs, ATPE gained an expanded range of applications utilizing physical phenomena occurring exclusively at the microscale. Today, research is being carried simultaneously in two different volume ranges, mL-scale (microtubes) and nL-scale (microchannels)...
September 14, 2016: Biotechnology Journal
Kousuke Moriyama, Shono Naito, Rie Wakabayashi, Masahiro Goto, Noriho Kamiya
Cellular spheroids have been received much attention in the biological and biomedical fields, especially as a base material for drug assays, regenerative medicine, and tissue engineering. Hydrogels have potential for scalable preparation of spheroids because they provide a three-dimensional environment suitable for three-dimensional cell cultivation. Herein, we report the potential use of a redox-responsive hydrogel as a scaffold for preparation and recovery of spheroids. A hydrogel composed of poly(ethylene glycol) (PEG), which can be degraded using cysteine as a reducing agent under mild conditions, was prepared by mixing an octa-thiolated PEG derivative (8-arm PEG-SH), horseradish peroxidase and a small phenolic compound (Glycyl-L-tyrosine)...
September 12, 2016: Biotechnology Journal
Jung Eun Yang, Je Woong Kim, Young Hoon Oh, So Young Choi, Hyuk Lee, A-Reum Park, Jihoon Shin, Si Jae Park, Sang Yup Lee
Polyhydroxyalkanoates (PHAs) containing 2-hydroxyacids such as lactate (LA) and 2-hydroxybutyrate (2HB) have recently been produced by metabolically engineered microorganisms. Here, we further expanded 2-hydroxyacid monomer spectrum of PHAs by engineering Escherichia coli to produce PHAs containing 2-hydroxyisovalerate (2HIV). To generate 2HIV in vivo, a feedback resistant ilvBN(mut) genes encoding acetohydroxyacid synthase and ilvCD genes encoding ketol-acid reductoisomerase and dihydroxyacid dehydratase, respectively, and panE gene encoding (D)-2-hydroxyacid dehydrogenase were overexpressed...
September 7, 2016: Biotechnology Journal
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