The circulation of products based on genetically modified (GM) organisms is highly regulated by some governments with strict implementation rules for the breeding, planting, marketing, labelling, and ...trading of such products. To ensure compliance, accurate detection methods for GM events are necessary, along with assurance that GM material falls within relevant threshold levels. The increasing complexity and potential of undocumented GM are a growing challenge for genetic screening. Here, we developed and assessed a highly multiplexed amplicon sequencing assay for the detection of GM events based on a microfluidics platform and next-generation sequencing (NGS). To probe GM events comprehensively, we designed a total of 230 new amplicons to cover flanking, promoter, junction, and coding sequences of GM sequences. In addition, we designed and implemented parallel amplification of ribosomal and chloroplast markers to define crop species identity from potentially mixed samples. Using reference GM material of 11 crop species and multiple amplicons, we successfully detected the presence of 10 known modifications per GM event. We also find that reported flanking sequences of GM events may not be all useful for diagnostic. We assessed the assay's potential to detect GM events in mixed samples as well as in highly diluted DNA. Finally, we performed a prospective search of potentially undocumented GM events in plant material. Our microfluidics-based amplicon GM detection approach fills important gaps in detecting potentially undocumented and complex GM events by recovering a wide range of specific amplicon sequences for evaluation. Integrating highly parallel amplicon assays in GM screening efforts should be an effective complement to aid post-market monitoring and regulatory compliance efforts.
•De novo amplicon design for consensus genetic modifications (GM) in plants.•Highly parallel amplicon sequencing across ∼200 samples based on Illumina sequencing.•Plant species identification of samples in parallel based on barcoding loci.•Robust distinction of GM material compared to control samples and with low input.•Novel next-generation sequencing framework to address future challenges of GMOs.
Atomically thin layers of two-dimensional materials can be assembled in vertical stacks that are held together by relatively weak van der Waals forces, enabling coupling between monolayer crystals ...with incommensurate lattices and arbitrary mutual rotation
. Consequently, an overarching periodicity emerges in the local atomic registry of the constituent crystal structures, which is known as a moiré superlattice
. In graphene/hexagonal boron nitride structures
, the presence of a moiré superlattice can lead to the observation of electronic minibands
, whereas in twisted graphene bilayers its effects are enhanced by interlayer resonant conditions, resulting in a superconductor-insulator transition at magic twist angles
. Here, using semiconducting heterostructures assembled from incommensurate molybdenum diselenide (MoSe
) and tungsten disulfide (WS
) monolayers, we demonstrate that excitonic bands can hybridize, resulting in a resonant enhancement of moiré superlattice effects. MoSe
and WS
were chosen for the near-degeneracy of their conduction-band edges, in order to promote the hybridization of intra- and interlayer excitons. Hybridization manifests through a pronounced exciton energy shift as a periodic function of the interlayer rotation angle, which occurs as hybridized excitons are formed by holes that reside in MoSe
binding to a twist-dependent superposition of electron states in the adjacent monolayers. For heterostructures in which the monolayer pairs are nearly aligned, resonant mixing of the electron states leads to pronounced effects of the geometrical moiré pattern of the heterostructure on the dispersion and optical spectra of the hybridized excitons. Our findings underpin strategies for band-structure engineering in semiconductor devices based on van der Waals heterostructures
.
Base editors (BEs) have been used to create C-to-T substitutions in various organisms. However, editing with rat APOBEC1-based BE3 is limited to a 5-nt sequence editing window and is inefficient in ...GC contexts. Here, we show that a base editor fusion protein composed of Cas9 nickase and human APOBEC3A (A3A-PBE) converts cytidine to thymidine efficiently in wheat, rice and potato with a 17-nucleotide editing window at all examined sites, independent of sequence context.
The valorization of byproducts from the sugarcane industry represents a potential alternative method with a low energy cost for the production of metabolites that are of commercial and industrial ...interest. The production of exopolysaccharides (EPSs) was carried out using the yeast Suhomyces kilbournensis isolated from agro-industrial sugarcane, and the products and byproducts of this agro-industrial sugarcane were used as carbon sources for their recovery. The effect of pH, temperature, and carbon and nitrogen sources and their concentration in EPS production by submerged fermentation (SmF) was studied in 170 mL glass containers of uniform geometry at 30 °C with an initial pH of 6.5. The resulting EPSs were characterized with Fourier-transform infrared spectroscopy (FT-IR). The results showed that the highest EPS production yields were 4.26 and 44.33 g/L after 6 h of fermentation using sucrose and molasses as carbon sources, respectively. Finally, an FT-IR analysis of the EPSs produced by S. kilbournensis corresponded to levan, corroborating its origin. It is important to mention that this is the first work that reports the production of levan using this yeast. This is relevant because, currently, most studies are focused on the use of recombinant and genetically modified microorganisms; in this scenario, Suhomyces kilbournensis is a native yeast isolated from the sugar production process, giving it a great advantage in the incorporation of carbon sources into their metabolic processes in order to produce levan sucrose, which uses fructose to polymerize levan.
Organoids in cancer research Drost, Jarno; Clevers, Hans
Nature reviews. Cancer,
07/2018, Letnik:
18, Številka:
7
Journal Article
Recenzirano
The recent advances in in vitro 3D culture technologies, such as organoids, have opened new avenues for the development of novel, more physiological human cancer models. Such preclinical models are ...essential for more efficient translation of basic cancer research into novel treatment regimens for patients with cancer. Wild-type organoids can be grown from embryonic and adult stem cells and display self-organizing capacities, phenocopying essential aspects of the organs they are derived from. Genetic modification of organoids allows disease modelling in a setting that approaches the physiological environment. Additionally, organoids can be grown with high efficiency from patient-derived healthy and tumour tissues, potentially enabling patient-specific drug testing and the development of individualized treatment regimens. In this Review, we evaluate tumour organoid protocols and how they can be utilized as an alternative model for cancer research.
Rapeseed (Brassica napus L.) is one of the most economically important oilseed crops worldwide. In Japan, it has been cultivated for more than a century and has formed many feral populations. The aim ...of this study was to elucidate the genetic diversity of feral rapeseeds by genotyping 537 individuals (among which 130 were determined to be genetically modified) sampled from various regions in Japan. Analysis of 30 microsatellite markers amplified 334 alleles and indicated moderate genetic diversity and high inbreeding (expected heterozygosity, 0.50; observed heterozygosity, 0.16; inbreeding coefficient within individuals, 0.68) within the feral populations. The Mantel test showed only an insignificant weak positive correlation between geographic distance and genetic distance. Analysis of molecular variance showed a greater genetic diversity among individuals than between populations. These results are in accordance with population structure assessed by using principal coordinate analysis and the program STRUCTURE, which showed that the 537 individuals could be assigned to 8 genetic clusters with very large genetic differences among individuals within the same geographic population, and that among feral individuals, many are closely related to rapeseed accessions in the NARO Genebank but some have unknown origins. These unique feral rapeseeds are likely to be affected by strong selection pressure. The results for genetically modified individuals also suggest that they have two different sources and have a considerable degree of diversity, which might be explained by hybridization with nearby individuals and separation of hybrid cultivars. The information obtained in this study could help improve the management of feral rapeseed plants in Japan.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Biohydrogen production, a feasible and environmentally friendly energy source, has advanced, notably in biocatalysts. This research carefully examines biocatalysts for biohydrogen synthesis and their ...recent advances. The debate begins with assessing biohydrogen's potential as an energy source and biocatalysts' importance in hydrogen production efficiency. Historical records reveal conventional methods and their limits, revealing novel microbial strains. The use of genetically modified organisms has altered hydrogen production by developing many methods and strains. Enzymatic mechanisms, especially novel enzymes and combinations, are crucial to hydrogen production. A multidisciplinary approach using microbiological, genetic, and enzymatic approaches can reveal synergistic effects and improve results. The techno-economic analysis compares the financial impacts and economic viability of these novel biocatalysts to conventional methods. Although there have been some improvements, scalability and safety issues remain unsolved. Future biocatalyst trends, developments, and wider implications for sustainable energy generation have been explored. The article examines the significance of these improvements and how biocatalysts can impact biohydrogen production.
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•Biocatalysts for enhanced biohydrogen production has been discussed.•Microbial engineering techniques for enhanced biohydrogen production has been elaborated.•Enzyme systems for enhanced biohydrogen production has been delineated.•Integrated systems for biohydrogen production have been explored.•Techno-economic analysis for biohydrogen production has been done.
Plant breeding plays a crucial role for the EU to live up to its values and promises of sustainability, innovation and diversity and inclusion. The current regulations, however, make it de facto ...impossible to use new breeding technologies.
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