@article{HaueisStechKubick2022,
  author    = {Haueis, Lisa and Stech, Marlitt and Kubick, Stefan},
  title     = {A Cell-free Expression Pipeline for the Generation and Functional Characterization of Nanobodies},
  series = {Frontiers in Bioengineering and Biotechnology},
  volume    = {10},
  journal   = {Frontiers in Bioengineering and Biotechnology},
  publisher = {Frontiers Media},
  address   = {Lausanne},
  issn      = {2296-4185},
  doi       = {10.3389/fbioe.2022.896763},
  pages     = {11},
  year      = {2022},
  abstract  = {Cell-free systems are well-established platforms for the rapid synthesis, screening, engineering and modification of all kinds of recombinant proteins ranging from membrane proteins to soluble proteins, enzymes and even toxins. Also within the antibody field the cell-free technology has gained considerable attention with respect to the clinical research pipeline including antibody discovery and production. Besides the classical full-length monoclonal antibodies (mAbs), so-called "nanobodies" (Nbs) have come into focus. A Nb is the smallest naturally-derived functional antibody fragment known and represents the variable domain (VHH, similar to 15 kDa) of a camelid heavy-chain-only antibody (HCAb). Based on their nanoscale and their special structure, Nbs display striking advantages concerning their production, but also their characteristics as binders, such as high stability, diversity, improved tissue penetration and reaching of cavity-like epitopes. The classical way to produce Nbs depends on the use of living cells as production host. Though cell-based production is well-established, it is still time-consuming, laborious and hardly amenable for high-throughput applications. Here, we present for the first time to our knowledge the synthesis of functional Nbs in a standardized mammalian cell-free system based on Chinese hamster ovary (CHO) cell lysates. Cell-free reactions were shown to be time-efficient and easy-to-handle allowing for the "on demand" synthesis of Nbs. Taken together, we complement available methods and demonstrate a promising new system for Nb selection and validation.},
  language  = {en}
}
@article{KrebsRakotoarinoroStechetal.2022,
  author    = {Krebs, Simon K. and Rakotoarinoro, Nathanael and Stech, Marlitt and Zemella, Anne and Kubick, Stefan},
  title     = {A CHO-based cell-free dual fluorescence reporter system for the straightforward assessment of amber suppression and scFv functionality},
  series = {Frontiers in Bioengineering and Biotechnology},
  volume    = {10},
  journal   = {Frontiers in Bioengineering and Biotechnology},
  publisher = {Frontiers Media},
  address   = {Lausanne},
  issn      = {2296-4185},
  doi       = {10.3389/fbioe.2022.873906},
  pages     = {15},
  year      = {2022},
  abstract  = {Incorporation of noncanonical amino acids (ncAAs) with bioorthogonal reactive groups by amber suppression allows the generation of synthetic proteins with desired novel properties. Such modified molecules are in high demand for basic research and therapeutic applications such as cancer treatment and in vivo imaging. The positioning of the ncAA-responsive codon within the protein's coding sequence is critical in order to maintain protein function, achieve high yields of ncAA-containing protein, and allow effective conjugation. Cell-free ncAA incorporation is of particular interest due to the open nature of cell-free systems and their concurrent ease of manipulation. In this study, we report a straightforward workflow to inquire ncAA positions in regard to incorporation efficiency and protein functionality in a Chinese hamster ovary (CHO) cell-free system. As a model, the well-established orthogonal translation components Escherichia coli tyrosyl-tRNA synthetase (TyrRS) and tRNATyr(CUA) were used to site-specifically incorporate the ncAA p-azido-l-phenylalanine (AzF) in response to UAG codons. A total of seven ncAA sites within an anti-epidermal growth factor receptor (EGFR) single-chain variable fragment (scFv) N-terminally fused to the red fluorescent protein mRFP1 and C-terminally fused to the green fluorescent protein sfGFP were investigated for ncAA incorporation efficiency and impact on antigen binding. The characterized cell-free dual fluorescence reporter system allows screening for ncAA incorporation sites with high incorporation efficiency that maintain protein activity. It is parallelizable, scalable, and easy to operate. We propose that the established CHO-based cell-free dual fluorescence reporter system can be of particular interest for the development of antibody-drug conjugates (ADCs).},
  language  = {en}
}
@article{CabukUenlue2022,
  author    = {{\c{C}}abuk, Uğur and {\"U}nl{\"u}, Ercan Sel{\c{c}}uk},
  title     = {A combined de novo assembly approach increases the quality of prokaryotic draft genomes},
  series = {Folia microbiologica : international journal for general, environmental and applied microbiology, and immunology},
  volume    = {67},
  journal   = {Folia microbiologica : international journal for general, environmental and applied microbiology, and immunology},
  publisher = {Springer},
  address   = {Dordrecht},
  issn      = {0015-5632},
  doi       = {10.1007/s12223-022-00980-7},
  pages     = {801 -- 810},
  year      = {2022},
  abstract  = {Next-generation sequencing methods provide comprehensive data for the analysis of structural and functional analysis of the genome. The draft genomes with low contig number and high N50 value can give insight into the structure of the genome as well as provide information on the annotation of the genome. In this study, we designed a pipeline that can be used to assemble prokaryotic draft genomes with low number of contigs and high N50 value. We aimed to use combination of two de novo assembly tools (SPAdes and IDBA-Hybrid) and evaluate the impact of this approach on the quality metrics of the assemblies. The followed pipeline was tested with the raw sequence data with short reads (< 300) for a total of 10 species from four different genera. To obtain the final draft genomes, we firstly assembled the sequences using SPAdes to find closely related organism using the extracted 16 s rRNA from it. IDBA-Hybrid assembler was used to obtain the second assembly data using the closely related organism genome. SPAdes assembler tool was implemented using the second assembly, produced by IDBA-hybrid as a hint. The results were evaluated using QUAST and BUSCO. The pipeline was successful for the reduction of the contig numbers and increasing the N50 statistical values in the draft genome assemblies while preserving the coverage of the draft genomes.},
  language  = {en}
}
@article{MbebiBreitlerBordeauxetal.2022,
  author    = {Mbebi, Alain J. and Breitler, Jean-Christophe and Bordeaux, M'elanie and Sulpice, Ronan and McHale, Marcus and Tong, Hao and Toniutti, Lucile and Castillo, Jonny Alonso and Bertrand, Benoit and Nikoloski, Zoran},
  title     = {A comparative analysis of genomic and phenomic predictions of growth-related traits in 3-way coffee hybrids},
  series = {G3: Genes, genomes, genetics},
  volume    = {12},
  journal   = {G3: Genes, genomes, genetics},
  number    = {9},
  publisher = {Genetics Soc. of America},
  address   = {Pittsburgh, PA},
  issn      = {2160-1836},
  doi       = {10.1093/g3journal/jkac170},
  pages     = {11},
  year      = {2022},
  abstract  = {Genomic prediction has revolutionized crop breeding despite remaining issues of transferability of models to unseen environmental conditions and environments. Usage of endophenotypes rather than genomic markers leads to the possibility of building phenomic prediction models that can account, in part, for this challenge. Here, we compare and contrast genomic prediction and phenomic prediction models for 3 growth-related traits, namely, leaf count, tree height, and trunk diameter, from 2 coffee 3-way hybrid populations exposed to a series of treatment-inducing environmental conditions. The models are based on 7 different statistical methods built with genomic markers and ChlF data used as predictors. This comparative analysis demonstrates that the best-performing phenomic prediction models show higher predictability than the best genomic prediction models for the considered traits and environments in the vast majority of comparisons within 3-way hybrid populations. In addition, we show that phenomic prediction models are transferrable between conditions but to a lower extent between populations and we conclude that chlorophyll a fluorescence data can serve as alternative predictors in statistical models of coffee hybrid performance. Future directions will explore their combination with other endophenotypes to further improve the prediction of growth-related traits for crops.},
  language  = {en}
}
@phdthesis{Gerling2022,
  author    = {Gerling, Marten Tobias},
  title     = {A microfluidic system for high-precision image-based live cell sorting using dielectrophoretic forces},
  doi       = {10.25932/publishup-58742},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-587421},
  school      = {Universit{\"a}t Potsdam},
  pages     = {vii, 87, VI},
  year      = {2022},
  abstract  = {An important goal in biotechnology and (bio-) medical research is the isolation of single cells from a heterogeneous cell population. These specialised cells are of great interest for bioproduction, diagnostics, drug development, (cancer) therapy and research. To tackle emerging questions, an ever finer differentiation between target cells and non-target cells is required. This precise differentiation is a challenge for a growing number of available methods. Since the physiological properties of the cells are closely linked to their morphology, it is beneficial to include their appearance in the sorting decision. For established methods, this represents a non addressable parameter, requiring new methods for the identification and isolation of target cells. Consequently, a variety of new flow-based methods have been developed and presented in recent years utilising 2D imaging data to identify target cells within a sample. As these methods aim for high throughput, the devices developed typically require highly complex fluid handling techniques, making them expensive while offering limited image quality. In this work, a new continuous flow system for image-based cell sorting was developed that uses dielectrophoresis to precisely handle cells in a microchannel. Dielectrophoretic forces are exerted by inhomogeneous alternating electric fields on polarisable particles (here: cells). In the present system, the electric fields can be switched on and off precisely and quickly by a signal generator. In addition to the resulting simple and effective cell handling, the system is characterised by the outstanding quality of the image data generated and its compatibility with standard microscopes. These aspects result in low complexity, making it both affordable and user-friendly. With the developed cell sorting system, cells could be sorted reliably and efficiently according to their cytosolic staining as well as morphological properties at different optical magnifications. The achieved purity of the target cell population was up to 95\% and about 85\% of the sorted cells could be recovered from the system. Good agreement was achieved between the results obtained and theoretical considerations. The achieved throughput of the system was up to 12,000 cells per hour. Cell viability studies indicated a high biocompatibility of the system. The results presented demonstrate the potential of image-based cell sorting using dielectrophoresis. The outstanding image quality and highly precise yet gentle handling of the cells set the system apart from other technologies. This results in enormous potential for processing valuable and sensitive cell samples.},
  language  = {en}
}
@article{FioriniDominguezReboredaetal.2022,
  author    = {Fiorini, Vanina D. and Dom{\´i}nguez, Marisol and Reboreda, Juan C. and Swaddle, John P.},
  title     = {A recent invasive population of the European starling sturnus vulgaris has lower genetic diversity and higher fluctuating asymmetry than primary invasive and native populations},
  series = {Biological invasions : unique international journal uniting scientists in the broad field of biological invasions},
  volume    = {24},
  journal   = {Biological invasions : unique international journal uniting scientists in the broad field of biological invasions},
  number    = {2},
  publisher = {Springer},
  address   = {Dordrecht},
  issn      = {1387-3547},
  doi       = {10.1007/s10530-021-02653-x},
  pages     = {437 -- 448},
  year      = {2022},
  abstract  = {Fluctuating asymmetries (FA) are small stress-induced random deviations from perfect symmetry that arise during the development of bilaterally symmetrical traits. One of the factors that can reduce developmental stability of the individuals and cause FA at a population level is the loss of genetic variation. Populations of founding colonists frequently have lower genetic variation than their ancestral populations that could be reflected in a higher level of FA. The European starling (Sturnus vulgaris) is native to Eurasia and was introduced successfully in the USA in 1890 and Argentina in 1983. In this study, we documented the genetic diversity and FA of starlings from England (ancestral population), USA (primary introduction) and Argentina (secondary introduction). We predicted the Argentinean starlings would have the highest level of FA and lowest genetic diversity of the three populations. We captured wild adult European starlings in England, USA, and Argentina, measured their mtDNA diversity and allowed them to molt under standardized conditions to evaluate their FA of primary feathers. For genetic analyses, we extracted DNA from blood samples of individuals from Argentina and USA and from feather samples from individuals from England and sequenced the mitochondrial control region. Starlings in Argentina showed the highest composite FA and exhibited the lowest haplotype and nucleotide diversity. The USA population showed a level of FA and genetic diversity similar to the native population. Therefore, the level of asymmetry and genetic diversity found among these populations was consistent with our predictions based on their invasion history.},
  language  = {en}
}
@misc{ApriyantoCompartFettke2022,
  author    = {Apriyanto, Ardha and Compart, Julia and Fettke, J{\"o}rg},
  title     = {A review of starch, a unique biopolymer - structure, metabolism and in planta modifications},
  series = {Plant science : an international journal of experimental plant biology},
  volume    = {318},
  journal   = {Plant science : an international journal of experimental plant biology},
  publisher = {Elsevier Science},
  address   = {Amsterdam [u.a.]},
  issn      = {0168-9452},
  doi       = {10.1016/j.plantsci.2022.111223},
  pages     = {8},
  year      = {2022},
  abstract  = {Starch is a complex carbohydrate polymer produced by plants and especially by crops in huge amounts. It consists of amylose and amylopectin, which have alpha-1,4-and alpha-1,6-linked glucose units. Despite this simple chemistry, the entire starch metabolism is complex, containing various (iso)enzymes/proteins. However, whose interplay is still not yet fully understood. Starch is essential for humans and animals as a source of nutrition and energy. Nowadays, starch is also commonly used in non-food industrial sectors for a variety of purposes. However, native starches do not always satisfy the needs of a wide range of (industrial) applications. This review summarizes the structural properties of starch, analytical methods for starch characterization, and in planta starch modifications.},
  language  = {en}
}
@article{PrueferWengerBieretal.2022,
  author    = {Pr{\"u}fer, Mareike and Wenger, Christian and Bier, Frank Fabian and Laux, Eva-Maria and H{\"o}lzel, Ralph},
  title     = {Activity of AC electrokinetically immobilized horseradish peroxidase},
  series = {Electrophoresis : microfluidics, nanoanalysis \& proteomics},
  journal   = {Electrophoresis : microfluidics, nanoanalysis \& proteomics},
  publisher = {Wiley},
  address   = {Hoboken},
  issn      = {0173-0835},
  doi       = {10.1002/elps.202200073},
  pages     = {1920 -- 1933},
  year      = {2022},
  abstract  = {Dielectrophoresis (DEP) is an AC electrokinetic effect mainly used to manipulate cells. Smaller particles, like virions, antibodies, enzymes, and even dye molecules can be immobilized by DEP as well. In principle, it was shown that enzymes are active after immobilization by DEP, but no quantification of the retained activity was reported so far. In this study, the activity of the enzyme horseradish peroxidase (HRP) is quantified after immobilization by DEP. For this, HRP is immobilized on regular arrays of titanium nitride ring electrodes of 500 nm diameter and 20 nm widths. The activity of HRP on the electrode chip is measured with a limit of detection of 60 fg HRP by observing the enzymatic turnover of Amplex Red and H2O2 to fluorescent resorufin by fluorescence microscopy. The initial activity of the permanently immobilized HRP equals up to 45\% of the activity that can be expected for an ideal monolayer of HRP molecules on all electrodes of the array. Localization of the immobilizate on the electrodes is accomplished by staining with the fluorescent product of the enzyme reaction. The high residual activity of enzymes after AC field induced immobilization shows the method's suitability for biosensing and research applications.},
  language  = {en}
}
@article{ReegStriglJeltsch2022,
  author    = {Reeg, Jette and Strigl, Lea and Jeltsch, Florian},
  title     = {Agricultural buffer zone thresholds to safeguard functional bee diversity},
  series = {Ecology and Evolution},
  volume    = {12},
  journal   = {Ecology and Evolution},
  edition   = {3},
  publisher = {Wiley Online Library},
  address   = {Hoboken, New Jersey, USA},
  issn      = {2045-7758},
  doi       = {10.1002/ece3.8748},
  pages     = {1 -- 17},
  year      = {2022},
  abstract  = {Wild bee species are important pollinators in agricultural landscapes. However, population decline was reported over the last decades and is still ongoing. While agricultural intensification is a major driver of the rapid loss of pollinating species, transition zones between arable fields and forest or grassland patches, i.e., agricultural buffer zones, are frequently mentioned as suitable mitigation measures to support wild bee populations and other pollinator species. Despite the reported general positive effect, it remains unclear which amount of buffer zones is needed to ensure a sustainable and permanent impact for enhancing bee diversity and abundance. To address this question at a pollinator community level, we implemented a process-based, spatially explicit simulation model of functional bee diversity dynamics in an agricultural landscape. More specifically, we introduced a variable amount of agricultural buffer zones (ABZs) at the transition of arable to grassland, or arable to forest patches to analyze the impact on bee functional diversity and functional richness. We focused our study on solitary bees in a typical agricultural area in the Northeast of Germany. Our results showed positive effects with at least 25\% of virtually implemented agricultural buffer zones. However, higher amounts of ABZs of at least 75\% should be considered to ensure a sufficient increase in Shannon diversity and decrease in quasi-extinction risks. These high amounts of ABZs represent effective conservation measures to safeguard the stability of pollination services provided by solitary bee species. As the model structure can be easily adapted to other mobile species in agricultural landscapes, our community approach offers the chance to compare the effectiveness of conservation measures also for other pollinator communities in future.},
  language  = {en}
}
@misc{ReegStriglJeltsch2022,
  author    = {Reeg, Jette and Strigl, Lea and Jeltsch, Florian},
  title     = {Agricultural buffer zone thresholds to safeguard functional bee diversity: Insights from a community modeling approach},
  series = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  journal   = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe},
  number    = {1281},
  issn      = {1866-8372},
  doi       = {10.25932/publishup-57080},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-570800},
  pages     = {17},
  year      = {2022},
  abstract  = {Wild bee species are important pollinators in agricultural landscapes. However, population decline was reported over the last decades and is still ongoing. While agricultural intensification is a major driver of the rapid loss of pollinating species, transition zones between arable fields and forest or grassland patches, i.e., agricultural buffer zones, are frequently mentioned as suitable mitigation measures to support wild bee populations and other pollinator species. Despite the reported general positive effect, it remains unclear which amount of buffer zones is needed to ensure a sustainable and permanent impact for enhancing bee diversity and abundance. To address this question at a pollinator community level, we implemented a process-based, spatially explicit simulation model of functional bee diversity dynamics in an agricultural landscape. More specifically, we introduced a variable amount of agricultural buffer zones (ABZs) at the transition of arable to grassland, or arable to forest patches to analyze the impact on bee functional diversity and functional richness. We focused our study on solitary bees in a typical agricultural area in the Northeast of Germany. Our results showed positive effects with at least 25\% of virtually implemented agricultural buffer zones. However, higher amounts of ABZs of at least 75\% should be considered to ensure a sufficient increase in Shannon diversity and decrease in quasi-extinction risks. These high amounts of ABZs represent effective conservation measures to safeguard the stability of pollination services provided by solitary bee species. As the model structure can be easily adapted to other mobile species in agricultural landscapes, our community approach offers the chance to compare the effectiveness of conservation measures also for other pollinator communities in future.},
  language  = {en}
}