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Abstract Microsatellite instability (MSI) occurs across a number of cancers and is associated with different clinical characteristics when compared to microsatellite stable (MSS) cancers. As MSI cancers have different characteristics, routine MSI testing is now recommended for a number of cancer types including colorectal cancer (CRC). Using gene panels for sequencing of known cancer mutations is routinely performed to guide treatment decisions. By adding a number of MSI regions to a small gene panel, the efficacy of simultaneous MSI detection in a series of CRCs was tested. Tumour DNA from formalin-fixed, paraffin-embedded (FFPE) tumours was sequenced using a 23-gene panel kit (ATOM-Seq) provided by GeneFirst. The mismatch repair (MMR) status was obtained for each patient from their routine pathology reports, and compared to MSI predictions from the sequencing data. By testing 29 microsatellite regions in 335 samples the MSI status was correctly classified in 314/319 samples (98.4% concordance), with sixteen failures. By reducing the number of regions in silico, comparable performance could be reached with as few as eight MSI marker positions. This test represents a quick, and accurate means of determining MSI status in FFPE CRC samples, as part of a routine gene mutation assay, and can easily be incorporated into a research or diagnostic setting. This could replace separate mutation and MSI tests with no loss of accuracy, thus improving testing efficiency.

Powdery mildew is one of the most problematic diseases in strawberry production. To date, few commercial strawberry cultivars are deemed to have complete resistance and as such, an extensive spray programme must be implemented to control the pathogen. Here, a large-scale field experiment was used to determine the powdery mildew resistance status of leaf and fruit tissues across a diverse panel of strawberry genotypes. This phenotypic data was used to identify Quantitative Trait Nucleotides (QTN) associated with tissue-specific powdery mildew resistance. In total, six stable QTN were found to be associated with foliar resistance, with one QTN on chromosome 7D associated with a 61% increase in resistance. In contrast to the foliage results, there were no QTN associated with fruit disease resistance and there was a high level of resistance observed on strawberry fruit, with no genetic correlation observed between fruit and foliar symptoms, indicating a tissue-specific response. Beyond the identification of genetic loci, we also demonstrate that genomic selection can lead to rapid gains in foliar resistance across genotypes, with the potential to capture >50% of the genetic foliage resistance present in the population. To date, breeding of robust powdery mildew resistance in strawberry has been impeded by the quantitative nature of natural resistance and a lack of knowledge relating to the genetic control of the trait. These results address this shortfall, through providing the community with a wealth of information that could be utilized for genomic informed breeding, implementation of which could deliver a natural resistance strategy for combatting powdery mildew.

Cherry (Prunus avium) fruits are important sources of vitamins, minerals, and nutrients in the human diet; however, they contain a large stone, making them inconvenient to eat ‘on the move’ and process. The exogenous application of gibberellic acid (GA3) can induce parthenocarpy in a variety of fruits during development. Here, we showed that the application of GA3 to sweet cherry unpollinated pistils acted as a trigger for fruit set and permitted the normal formation of fruit up to a period of twenty-eight days, indicating that gibberellins are involved in the activation of the cell cycle in the ovary wall cells, leading to fruit initiation. However, after this period, fruit development ceased and developing fruit began to be excised from the branch by 35 days post treatment. This work also showed that additional signals are required for the continued development of fully mature parthenocarpic fruit in sweet cherry.

Abstract This study aims to establish whether zinc (Zn) and cadmium (Cd) share similar physiological mechanisms for uptake and translocation in cacao plants (Theobroma cacao L.). Multiple-collector ICP-MS was used to determine the Zn stable isotope compositions in the roots, stems and leaves of 19 diverse cacao genotypes grown in hydroponics with 20 µmol L−1 CdCl2. Additional plants of one genotype were grown in hydroponic solutions containing lower Cd concentrations (0 and 5 µmol L−1 added CdCl2). Regardless of the Cd concentration used in the exposures, the Zn stable isotope compositions show the same systematic patterns in plant organs, with δ66Znroot > δ66Znstem > δ66Znleaf (δ66Zn denotes relative differences in 66Zn/64Zn ratios in parts per thousand). The mean Zn stable isotope fractionation between the plants and the hydroponic solutions was ε66Znuptake = –1.15 ± 0.36‰ (2SD), indicating preferential uptake of isotopically light Zn by plants from the hydroponic solution. The mean stable isotope fractionation factor associated with translocation of Zn from roots to shoots, ε66Znseq-mob = + 0.52 ± 0.36‰ (2SD), shows that isotopically heavy Zn is preferentially sequestered in the cacao roots, whilst isotopically light Zn is mobilised to the leaves. A comparison with the Cd stable isotope compositions of the same plants shows that both isotopically light Zn and Cd are preferentially taken up by cacao plants. In contrast to Zn, however, the cacao roots retain isotopically light Cd and transfer isotopically heavy Cd to the leaves.

Abstract Through agronomic traits and sequencing data, the cultivated and wild varieties of grapes and peaches were analyzed and compared in terms of fruit size, fruit flavor, fruit resistance, and fruit color. Cultivated grapes and peaches have advantages in fruit size, soluble sugar content, sugar and acid ratio, etc. Wild grapes and peaches have utility value in resistance. The results showed that there were 878 and 301 differentially expressed genes in cultivated and wild grapes and peaches in the three growth stages, respectively based on the next-generation sequencing study. Ten and twelve genes related to the differences between cultivated and wild grapes and peaches were found respectively. Among them, three genes, namely chalcone synthase (CHS), glutathione S-transferase (GST) and malate dehydrogenase (MDH1) were present in both cultivated and wild grapes and peaches.

High concentrations of toxic cadmium (Cd) in soils are problematic as the element accumulates in food crops such as rice and cacao. A mitigation strategy to minimise Cd accumulation is to enhance the competitive uptake of plant-essential metals. Theobroma cacao seedlings were grown hydroponically with added Cd. Eight different treatments were used, which included/excluded hydroponic or foliar zinc (Zn) and/or iron (Fe) for the final growth period. Analyses of Cd concentrations and natural stable isotope compositions by multiple collector ICP-MS were conducted. Cadmium uptake and translocation decreased when Fe was removed from the hydroponic solutions, while the application of foliar Zn-EDTA may enhance Cd translocation. No significant differences in isotope fractionation during uptake were found between treatments. Data from all treatments fit a single Cd isotope fractionation model associated with sequestration (seq) of isotopically light Cd in roots and unidirectional mobilisation (mob) of isotopically heavier Cd to the leaves (ε114Cdseq-mob = −0.13‰). This result is in excellent agreement with data from an investigation of 19 genetically diverse cacao clones. The different Cd dynamics exhibited by the clones and seen in response to different Fe availability may be linked to similar physiological processes, such as the regulation of specific transporter proteins.

Metrics seeking to predict financial risk-taking behaviors typicallyexhibit limited validity. This is due to the fluid nature of anindividual’s risk taking, and the influence of the mode and medium,which presents a decision. This paper presents two experiments thatinvestigate how an existing risk elicitation task’s predictive capacitymay be enhanced through the application of an interactive model ofvisual reasoning in a digitized version. In the first experiment, 60participants demonstrated their reasoning process. In the secondexperiment, 225 participants were randomly assigned into threegroups, with the validated risk elicitation task compared as a controlto interactive digital and non-interactive digital stimuli with piecharts. The experiments yielded significant results, highlighting thatwhen participants interact with a graph to reason their choices, itleads to consistent choices. The findings have implications forimprovement of the risk task's validity and the deployment of digitalinteractive assessments beyond laboratory settings.

Abstract Background European canker, caused by the fungal pathogen Neonectria ditissima, is an economically damaging disease in apple producing regions of the world – especially in areas with moderate temperatures and high rainfall. The pathogen has a wide host range of hardwood perennial species, causing trunk cankers, dieback and branch lesions in its hosts. Although apple scion germplasm carrying partial resistance to the disease has been described, little is still known of the genetic basis for this quantitative resistance. Results Resistance to Neonectria ditissima was studied in a multiparental population of apple scions using several phenotyping methods. The studied population consists of individuals from multiple families connected through a common pedigree. The degree of disease of each individual in the population was assessed in three experiments: artificial inoculations of detached dormant shoots, potted trees in a glasshouse and in a replicated field experiment. The genetic basis of the differences in disease was studied using a pedigree-based analysis (PBA). Three quantitative trait loci (QTL), on linkage groups (LG) 6, 8 and 10 were identified in more than one of the phenotyping strategies. An additional four QTL, on LG 2, 5, 15 and 16 were only identified in the field experiment. The QTL on LG2 and 16 were further validated in a biparental population. QTL effect sizes were small to moderate with 4.3 to 19% of variance explained by a single QTL. A subsequent analysis of QTL haplotypes revealed a dynamic response to this disease, in which the estimated effect of a haplotype varied over the field time-points. Conclusions This study describes the first identified QTL associated with resistance to N. ditissima in apple scion germplasm. The results from this study show that QTL present in germplasm commonly used in apple breeding have a low to medium effect on resistance to N. ditissima. Hence, multiple QTL will need to be considered to improve resistance through breeding.

Abstract In contrast to most land plant species, sorbitol, instead of sucrose, is the major photosynthetic product in many Rosaceae species. It has been well illustrated that three key functional genes encoding sorbitol-6-phosphate dehydrogenase (S6PDH), sorbitol dehydrogenase (SDH), and sorbitol transporter (SOT), are mainly responsible for the synthesis, degradation and transportation of sorbitol. In this study, the genome-wide identification of S6PDH, SDH and SOT genes was conducted in four Rosaceae species, peach, mei, apple and pear, and showed the sorbitol bio-pathway to be dominant (named sorbitol present group, SPG); another three related species, including tomato, poplar and Arabidopsis, showed a non-sorbitol bio-pathway (named sorbitol absent group, SAG). To understand the evolutionary differences of the three important gene families between SAG and SPG, their corresponding gene duplication, evolutionary rate, codon bias and positive selection patterns have been analyzed and compared. The sorbitol pathway genes in SPG were found to be expanded through dispersed and tandem gene duplications. Branch-specific model analyses revealed SDH and S6PDH clade A were under stronger purifying selection in SPG. A higher frequency of optimal codons was found in S6PDH and SDH than that of SOT in SPG, confirming the purifying selection effect on them. In addition, branch-site model analyses revealed SOT genes were under positive selection in SPG. Expression analyses showed diverse expression patterns of sorbitol-related genes. Overall, these findings provide new insights in the evolutionary characteristics for the three key sorbitol metabolism-related gene families in Rosaceae and other non-sorbitol dominant pathway species.

This study employed a phenomenographic approach to investigate science teachers' conceptions of inquiry-based learning through a serious game. Simaula is a prototype game designed and used as a virtual practicum for eliciting understandings on how in-game inquiry was appeared to, or experienced by, the participating teachers. Group interviews with 20 secondary education science teachers revealed four qualitatively different ways of experiencing inquiry-based learning through Simaula: (a) as uncovering insights about student's learning needs, interests and emotions; (b) as generating ideas and concepts for meaningful inquiry; (c) as a set of operations for designing and carrying out scientific research; and (d) as authentic inquiry for enabling knowledge building processes. Seven dimensions of variation have been identified viewed as contextual influences on conceptions of in-game inquiry constituting discernment of: epistemic inquiry-based learning modes; role of teacher; role of student; game-play focus; core mechanics focus; feedback and progress mechanics and game uncertainty. The results illuminated a partial in-game inquiry approach with distinct epistemic modes from developing empathy and meaning making to knowledge construction and knowledge building. The findings also indicated that game design elements played central role in shaping conceptions of in-game inquiry from focusing on rules and logic as means to completing the game's level to understanding the complexity of core mechanics for developing and transferring in-game inquiry to the real classroom. This insinuates that distinct game design properties may be considered in terms of extending intrinsic in-game inquiry experiences to actual in-class inquiry practice.

The cocoa crop (Theobroma cacao L.) is known to be a host for several badnaviruses, some of which cause severe disease, while others are asymptomatic. Recently, the first preliminary evidence has been published concerning the occurrence of a polerovirus in cacao. We report here the first near-complete genome sequence of cacao polerovirus (CaPV) by combining bioinformatic searches of cacao transcript databases, with cloning from the infected germplasm. The reported novel genome has all the genome features known for poleroviruses from other species. Pairwise identity analyses of RNA-dependent RNA polymerase and coat protein indicates < 60% similarity of CaPV with any reported poleroviruses; hence, we propose that the polerovirus isolate reported in this study is a novel polerovirus. The genome sequence information was also used to develop a multiplex RT-PCR assay, which was applied to screen a selected range of germplasms and to identify several infected clones. Although there is no evidence that this virus causes any severe disease, this new information, together with a robust diagnostic assay, are of strategic importance in developing protocols for the safe international transfer of cacao germplasms.

To address domestication and improvement studies of soybean seed size- and oil-related traits, a series of domesticated and improved regions, loci, and candidate genes were identified in 286 soybean accessions using domestication and improvement analyses, genome-wide association studies, quantitative trait locus (QTL) mapping and bulked segregant analyses in this study. As a result, 534 candidate domestication regions (CDRs) and 458 candidate improvement regions (CIRs) were identified in this study and integrated with those in five and three previous studies, respectively, to obtain 952 CDRs and 538 CIRs; 1469 loci for soybean seed size- and oil-related traits were identified in this study and integrated with those in Soybase to obtain 433 QTL clusters. The two results were intersected to obtain 245 domestication and 221 improvement loci for the above traits. Around these trait-related domestication and improvement loci, 7 domestication and 7 improvement genes were found to be truly associated with these traits, and 372 candidate domestication and 87 candidate improvement genes were identified using gene expression, SNP variants in genome, miRNA binding, KEGG pathway, DNA methylation, and haplotype analysis. These genes were used to explain the trait changes in domestication and improvement. As a result, the trait changes can be explained by their frequencies of elite haplotypes, base mutations in coding region, and three factors affecting their expression levels. In addition, 56 domestication and 15 improvement genes may be valuable for future soybean breeding. This study can provide useful gene resources for future soybean breeding and molecular biology research.

Abstract Background Pears and apples are both perennial deciduous trees of the Rosaceae family, and both are important economic fruit trees worldwide. The emergence of many varieties in the market has been mostly domesticated from wild to cultivated and regulated by the differential expression of genes. However, the molecular process and pathways underlying this phenomenon remain unclear. Four typical wild and cultivar pear and apple trees at three developmental stages were used in our study to investigate the molecular process at the transcriptome level. Result Physiological observations indicated the obvious differences of size, weight, sugar acid content and peel color in wild and cultivar fruit among each developmental stage. Using next-generation sequencing based RNA-seq expression profiling technology, we produced a transcriptome in procession of a large fraction of annotated pear and apple genes, and provided a molecular basis underlying the phenomenon of wild and cultivar fruit tree differences. 5921 and 5744 differential expression genes were identified in pear and apple at three developmental stages respectively. We performed temporal and spatial differential gene expression profiling in developing fruits. Several key pathways such as signal transduction, photosynthesis, translation and many metabolisms were identified as involved in the differentiation of wild and cultivar fruits. Conclusion In this study, we reported on the next-generation sequencing study of the temporal and spatial mRNA expression profiling of pear and apple fruit trees. Also, we demonstrated that the integrated analysis of pear and apple transcriptome, which strongly revealed the consistent process of domestication in Rosaceae fruit trees. The results will be great influence to the improvement of cultivar species and the utilization of wild resources.

Abstract This study employed a phenomenographic approach to investigate science teachers’ conceptions of inquiry-based learning through a serious game. Simaula is a prototype game designed and used as a virtual practicum for eliciting understandings on how in-game inquiry was appeared to, or experienced by, the participating teachers. Group interviews with 20 secondary education science teachers revealed four qualitatively different ways of experiencing inquiry-based learning through Simaula: (a) as uncovering insights about student’s learning needs, interests and emotions; (b) as generating ideas and concepts for meaningful inquiry; (c) as a set of operations for designing and carrying out scientific research; and (d) as authentic inquiry for enabling knowledge building processes. Seven dimensions of variation have been identified viewed as contextual influences on conceptions of in-game inquiry constituting discernment of: epistemic inquiry-based learning modes; role of teacher; role of student; game-play focus; core mechanics focus; feedback and progress mechanics and game uncertainty. The results illuminated a partial in-game inquiry approach with distinct epistemic modes from developing empathy and meaning making to knowledge construction and knowledge building. The findings also indicated that game design elements played central role in shaping conceptions of in-game inquiry from focusing on rules and logic as means to completing the game’s level to understanding the complexity of core mechanics for developing and transferring in-game inquiry to the real classroom. This insinuates that distinct game design properties may be considered in terms of extending intrinsic in-game inquiry experiences to actual in-class inquiry practice.

Abstract Theobroma cacao is one of the most economically important tropical trees, being the source of chocolate. As part of an ongoing study to understand the diversity of the badnavirus complex, responsible for the cacao swollen shoot virus disease in West Africa, evidence was found recently of virus-like sequences in asymptomatic cacao plants. The present study exploited the wealth of genomic resources in this crop, and combined bioinformatic, molecular, and genetic approaches to report for the first time the presence of integrated badnaviral sequences in most of the cacao genetic groups. These sequences, which we propose to name eTcBV for endogenous T. cacao bacilliform virus, varied in type with each predominating in a specific genetic group. A diagnostic multiplex PCR method was developed to identify the homozygous or hemizygous condition of one specific insert, which was inherited as a single Mendelian trait. These data suggest that these integration events occurred before or during the species diversification in Central and South America, and prior to its cultivation in other regions. Such evidence of integrated sequences is relevant to the management of cacao quarantine facilities and may also aid novel methods to reduce the impact of such viruses in this crop.

Lignin is one of the main components of plant cell walls, which provides mechanical support for plants and also contributes to resisting against plant pathogenic fungi. In the fruit industry, the lignin content can affect the quality of fruit. The biosynthesis of lignin involves a variety of enzymes, of which caffeoyl-CoA 3-O-methyltransferase (CCoAOMT) is a class of methyltransferases that plays an essential role in lignin biosynthesis. Studies have been conducted on the CCoAOMT gene family in several species, including arabidopsis (Arabidopsis thaliana), black poplar (Populus nigra), and cotton (Gossypium hirsutum). Still, there is relatively little research on this gene family in the Rosaceae. In this study, we used bioinformatics to identify and characterize the CCoAOMT gene family in apple (Malus domestica), chinese white pear (Pyrus bretschneideri), and peach (Prunus persica). In total, 35 CCoAOMT genes were identified in the three Rosaceae species: 8 from chinese white pear, 12 from apple, and 15 from peach. By using structure analysis and collinearity analysis, we found 12 conserved motifs and 12 pairs of CCoAOMT genes with collinearity. In the phylogenetic tree, the gene family was mainly divided into two groups. The genes had different expression patterns during the growth and development stage of fruit, a finding that is consistent with the pattern of lignin accumulation. This study will be beneficial for further study of CCoAOMT genes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Abstract Liquid biopsy testing utilising Next Generation Sequencing (NGS) is rapidly moving towards clinical adoption for personalised oncology. However, before NGS can fulfil its potential any novel testing approach must identify ways of reducing errors, allowing separation of true low-frequency mutations from procedural artefacts, and be designed to improve upon current technologies. Popular NGS technologies typically utilise two DNA capture approaches; PCR and ligation, which have known limitations and seem to have reached a development plateau with only small, stepwise improvements being made. To maximise the ultimate utility of liquid biopsy testing we have developed a highly versatile approach to NGS: Adaptor Template Oligo Mediated Sequencing (ATOM-Seq). ATOM-Seq's strengths and versatility avoid the major limitations of both PCR- and ligation-based approaches. This technology is ligation free, simple, efficient, flexible, and streamlined, and it offers novel advantages that make it perfectly suited for use on highly challenging clinical material. Using reference and clinical materials, we demonstrate detection of known SNVs down to allele frequencies of 0.1% using as little as 20–25 ng of cfDNA, as well as the ability to detect fusions from RNA. We illustrate ATOM-Seq’s suitability for clinical testing by showing high concordance rates between paired cfDNA and FFPE clinical samples.

We report that the mammalian 5-methylcytosine (5mC) oxidase Tet3 exists as three major isoforms and characterized the full-length isoform containing an N-terminal CXXC domain (Tet3FL). This CXXC domain binds to unmethylated CpGs, but, unexpectedly, its highest affinity is toward 5-carboxylcytosine (5caC). We determined the crystal structure of the CXXC domain-5caC-DNA complex, revealing the structural basis of the binding specificity of this domain as a reader of CcaCG sequences. Mapping of Tet3FL in neuronal cells shows that Tet3FL is localized precisely at the transcription start sites (TSSs) of genes involved in lysosome function, mRNA processing, and key genes of the base excision repair pathway. Therefore, Tet3FL may function as a regulator of 5caC removal by base excision repair. Active removal of accumulating 5mC from the TSSs of genes coding for lysosomal proteins by Tet3FL in postmitotic neurons of the brain may be important for preventing neurodegenerative diseases.

The methodologies and software packages for mapping quantitative trait loci (QTLs) in bi-parental segregation populations are well established. However, it is still difficult to detect small-effect and linked QTLs. To address this issue, we proposed a genome-wide composite interval mapping (GCIM) in bi-parental segregation populations. To popularize this method, we developed an R package. This program with two versions (Graphical User Interface: QTL.gCIMapping.GUI v2.0 and code: QTL.gCIMapping v3.2) can be used to identify QTLs for quantitative traits in recombinant inbred lines, doubled haploid lines, backcross and F2 populations. To save running time, fread function was used to read the dataset, parallel operation was used in parameter estimation, and conditional probability calculation was implemented by C++. Once one input file with *.csv or *.txt formats is uploaded into the package, one or two output files and one figure can be obtained. The input file with the ICIM and win QTL cartographer formats is available as well. Real data analysis for 1000-grain weight in rice showed that the GCIM detects the maximum previously reported QTLs and genes, and has the minimum AIC value in the stepwise regression of all the identified QTLs for this trait; using stepwise regression and empirical Bayesian analyses, there are some false QTLs around the previously reported QTLs and genes from the CIM method. The above software packages on Windows, Mac and Linux can be downloaded from https://cran.r-project.org/web/packages/ or https://bigd.big.ac.cn/biocode/tools/7078/releases/27 in order to identify all kinds of omics QTLs.

Somatic embryogenesis (SE), which occurs naturally in many plant species, serves as a model to elucidate cellular and molecular mechanisms of embryo patterning in plants. Decoding the regulatory landscape of SE is essential for its further application. Hence, the present study was aimed at employing Weighted Gene Correlation Network Analysis (WGCNA) to construct a gene coexpression network (GCN) for Arabidopsis SE and then identifying highly correlated gene modules to uncover the hub genes associated with SE that may serve as potential molecular targets. A total of 17,059 genes were filtered from a microarray dataset comprising four stages of SE, i.e., stage I (zygotic embryos), stage II (proliferating tissues at 7 days of induction), stage III (proliferating tissues at 14 days of induction), and stage IV (mature somatic embryos). This included 1,711 transcription factors and 445 EMBRYO DEFECTIVE genes. GCN analysis identified a total of 26 gene modules with the module size ranging from 35 to 3,418 genes using a dynamic cut tree algorithm. The module-trait analysis revealed that four, four, seven, and four modules were associated with stages I, II, III, and IV, respectively. Further, we identified a total of 260 hub genes based on the degree of intramodular connectivity. Validation of the hub genes using publicly available expression datasets demonstrated that at least 78 hub genes are potentially associated with embryogenesis; of these, many genes remain functionally uncharacterized thus far. In silico promoter analysis of these genes revealed the presence of cis-acting regulatory elements, “soybean embryo factor 4 (SEF4) binding site,” and “E-box” of the napA storage-protein gene of Brassica napus; this suggests that these genes may play important roles in plant embryo development. The present study successfully applied WGCNA to construct a GCN for SE in Arabidopsis and identified hub genes involved in the development of somatic embryos. These hub genes could be used as molecular targets to further elucidate the molecular mechanisms underlying SE in plants.

Feedback is a critical aspect of optimised learning design, but there are few, if any, feedback models that map different types of feedback and how they may assist students to increase performance and enhance their learning experience. This research paper outlines a feedback model as an extension of the four-dimensional framework which includes a consideration of the type, the content, the format, and the frequency of feedback, as well as the agent which delivers it. This model is based upon an understanding of learning in the context of designing learning experiences and utilises a game-based model of learning to understand the importance of motivation and autonomy in learners to enhance and accelerate learning. The framework is developed and reflected upon by analysing two cases: a medical triage case in which the timing and frequency of feedback proved critical, and a business simulation which illuminated the need for a range of types of feedback and to be aware of the possibility of different agents (instructor peer and game) that can deliver feedback. The extended model may help game and learning designers alike to discern different types of feedback, both in games and more generally, in more explicit and nuanced ways.

Variation in patterns of gene expression can result from modifications in the genome that occur without a change in the sequence of the DNA; such modifications include methylation of cytosine to generate 5-methylcytosine (5mC) resulting in the generation of heritable epimutation and novel epialleles. This type of non-sequence variation is called epigenetics. The enzymes responsible for generation of such DNA modifications in mammals are named DNA methyltransferases (DNMT) including DNMT1, DNMT2 and DNMT3. The later stages of oxidations to these modifications are catalyzed by Ten Eleven Translocation (TET) proteins, which contain catalytic domains belonging to the 2-oxoglutarate dependent dioxygenase family. In various mammalian cells/tissues including embryonic stem cells, cancer cells and brain tissues, it has been confirmed that these proteins are able to induce the stepwise oxidization of 5-methyl cytosine to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and finally 5-carboxylcytosine (5caC). Each stage from initial methylation until the end of the DNA demethylation process is considered as a specific epigenetic mark that may regulate gene expression. This review discusses controversial evidence for the presence of such oxidative products, particularly 5hmC, in various plant species. Whereas some reports suggest no evidence for enzymatic DNA demethylation, other reports suggest that the presence of oxidative products is followed by the active demethylation and indicate the contribution of possible TET-like proteins in the regulation of gene expression in plants. The review also summarizes the results obtained by expressing the human TET conserved catalytic domain in transgenic plants.

This paper consolidates evidence and material from a range of specialist and disciplinary fields to provide an evidence-based review and synthesis on the design and use of serious games in higher education. Search terms identified 165 papers reporting conceptual and empirical evidence on how learning attributes and game mechanics may be planned, designed and implemented by university teachers interested in using games, which are integrated into lesson plans and orchestrated as part of a learning sequence at any scale. The findings outline the potential of classifying the links between learning attributes and game mechanics as a means to scaffold teachers' understanding of how to perpetuate learning in optimal ways while enhancing the in-game learning experience. The findings of this paper provide a foundation for describing methods, frames and discourse around experiences of design and use of serious games, linked to methodological limitations and recommendations for further research in this area.

Fruits are an important source of vitamins, minerals and nutrients in the human diet. They also contain several compounds of nutraceutical importance that have significant antioxidant and anti-inflammatory roles, which can protect the consumer from diseases, such as cancer, and cardiovascular disease as well as having roles in reducing the build-up of LDL-cholesterol in blood plasma and generally reduce the risks of disease and age-related decline in health. Cherries contain high concentrations of bioactive compounds and minerals, including calcium, phosphorous, potassium and magnesium, and it is, therefore, unsurprising that cherry consumption has a positive impact on health. This review highlights the development of sweet cherry fruit, the health benefits of cherry consumption, and the options for increasing consumer acceptance and consumption.

The oomycete Phytophthora fragariae is a highly destructive pathogen of cultivated strawberry (Fragaria × ananassa), causing the root rotting disease, “red core”. The host-pathogen interaction has a well described gene-for-gene resistance relationship, but to date neither candidate avirulence nor resistance genes have been identified. We sequenced a set of American, Canadian, and United Kingdom isolates of known race type, along with three representatives of the closely related pathogen of the raspberry (Rubus idaeus), P. rubi, and found a clear population structure, with a high degree of nucleotide divergence seen between some race types and abundant private variation associated with race types 4 and 5. In contrast, between isolates defined as United Kingdom races 1, 2, and 3 (UK1-2-3) there was no evidence of gene loss or gain; or the presence of insertions/deletions (INDELs) or Single Nucleotide Polymorphisms (SNPs) within or in proximity to putative pathogenicity genes could be found associated with race variation. Transcriptomic analysis of representative UK1-2-3 isolates revealed abundant expression variation in key effector family genes associated with pathogen race; however, further long read sequencing did not reveal any long range polymorphisms to be associated with avirulence to race UK2 or UK3 resistance, suggesting either control in trans or other stable forms of epigenetic modification modulating gene expression. This work reveals the combined power of population resequencing to uncover race structure in pathosystems and in planta transcriptomic analysis to identify candidate avirulence genes. This work has implications for the identification of putative avirulence genes in the absence of associated expression data and points toward the need for detailed molecular characterisation of mechanisms of effector regulation and silencing in oomycete plant pathogens.

L-Ascorbic acid (ascorbate, Vitamin C) is an essential human micronutrient that is predominantly obtained from plants. It is known to work as the major antioxidant in plants, and it underpins several environmentally induced stresses due to its use as a co-factor by certain 2-oxoglutarate-dependent (2-OG) dioxygenases [2(OG)-dioxygenases]. It is important to understand the role of 2(OG)-dioxygenases in the biosynthesis of ascorbate. The present study examined contents of ascorbate and protein-protein interaction in nine T-DNA mutants of Arabidopsis containing an insert in their respective (2-OG) dioxygenase genes (At1g20270, At1g68080, At2g17720, At3g06290, At3g28490, At4g35810, At4g35820, At5g18900, At5g66060). In this study, the amount of ascorbate in five of the mutants was shown to be almost two-fold or more than two-fold higher than in the wild type. This result may be a consequence of the insertion of the T-DNA. The prediction of possible protein interactions between 2(OG)-dioxygenases and relevant ascorbate-function players may indicate the oxidative effects of certain dioxygenase proteins in plants. It is expected that certain dioxygenases are actively involved in the metabolic and biosynthetic pathways of ascorbate. This involvement may be of importance to increase ascorbate amounts in plants for human nutrition, and to protect plant species against stress conditions.

Abstract Background Rapeseed (Brassica napus L.) and soybean (Glycine max L.) seeds are rich in both protein and oil, which are major sources of biofuels and nutrition. Although the difference in seed oil content between soybean (~ 20%) and rapeseed (~ 40%) exists, little is known about its underlying molecular mechanism. Results An integrated omics analysis was performed in soybean, rapeseed, Arabidopsis (Arabidopsis thaliana L. Heynh), and sesame (Sesamum indicum L.), based on Arabidopsis acyl-lipid metabolism- and carbon metabolism-related genes. As a result, candidate genes and their transcription factors and microRNAs, along with phylogenetic analysis and co-expression network analysis of the PEPC gene family, were found to be largely associated with the difference between the two species. First, three soybean genes (Glyma.13G148600, Glyma.13G207900 and Glyma.12G122900) co-expressed with GmPEPC1 are specifically enriched during seed storage protein accumulation stages, while the expression of BnPEPC1 is putatively inhibited by bna-miR169, and two genes BnSTKA and BnCKII are co-expressed with BnPEPC1 and are specifically associated with plant circadian rhythm, which are related to seed oil biosynthesis. Then, in de novo fatty acid synthesis there are rapeseed-specific genes encoding subunits β-CT (BnaC05g37990D) and BCCP1 (BnaA03g06000D) of heterogeneous ACCase, which could interfere with synthesis rate, and β-CT is positively regulated by four transcription factors (BnaA01g37250D, BnaA02g26190D, BnaC01g01040D and BnaC07g21470D). In triglyceride synthesis, GmLPAAT2 is putatively inhibited by three miRNAs (gma-miR171, gma-miR1516 and gma-miR5775). Finally, in rapeseed there was evidence for the expansion of gene families, CALO, OBO and STERO, related to lipid storage, and the contraction of gene families, LOX, LAH and HSI2, related to oil degradation. Conclusions The molecular mechanisms associated with differences in seed oil content provide the basis for future breeding efforts to improve seed oil content.

Abstract Background ETCHbox genes are eutherian-specific homeobox genes expressed during preimplantation development at a time when the first cell lineage decisions are being made. The mouse has an unusual repertoire of ETCHbox genes with several gene families lost in evolution and the remaining two, Crxos and Obox, greatly divergent in sequence and number. Each has undergone duplication to give a double homeodomain Crxos locus and a large cluster of over 60 Obox loci. The gene content differences between species raise important questions about how evolution can tolerate loss of genes implicated in key developmental events. Results We find that Crxos internal duplication occurred in the mouse lineage, while Obox duplication was stepwise, generating subgroups with distinct sequence and expression. Ectopic expression of three Obox genes and a Crxos transcript in primary mouse embryonic cells followed by transcriptome sequencing allowed investigation into their functional roles. We find distinct transcriptomic influences for different Obox subgroups and Crxos, including modulation of genes related to zygotic genome activation and preparation for blastocyst formation. Comparison with similar experiments performed using human homeobox genes reveals striking overlap between genes downstream of mouse Crxos and genes downstream of human ARGFX. Conclusions Mouse Crxos and human ARGFX homeobox genes are paralogous rather than orthologous, yet they have evolved to regulate a common set of genes. This suggests there was compensation of function alongside gene loss through co-option of a different locus. Functional compensation by non-orthologous genes with dissimilar sequences is unusual but may indicate underlying distributed robustness. Compensation may be driven by the strong evolutionary pressure for successful early embryo development.

Cocoa, Theobroma cacao, is an important tropical perennial crop grown widely in the humid tropics. The exchange of cocoa germplasm between germplasm collections and breeding centres is vital for varietal development. Intermediate quarantine facilities, such as the International Cocoa Quarantine Centre, Reading UK (ICQC-R) play a vital role in ensuring the transfer of germplasm whilst minimising the risk of spreading pests and diseases. Current screening procedures combine visual inspection and molecular techniques, which are effective in detecting Cocoa swollen shoot virus (CSSV), a badnavirus, which causes severe losses but are restricted to West Africa. However, the detection of latent or mild virus infections that produce no visual symptoms has been a challenge. Recently two badnavirus species of cocoa producing mild symptoms, cacao mild mosaic virus (CaMMV) and cacao yellow vein-banding virus (CYVBV), have been sequenced. Here, we report new assays for the detection of these two species, for the first time in non-symptomatic accessions. Evolutionary and bioinformatic analyses of the viruses suggest their most recent source was from Trinidad, though there is historic evidence that these viruses may have their origin in South America and then become widespread globally over the last century. We also report a novel colorimetric Loop-mediated isothermal amplification (LAMP) assay for the detection of CYVBV. This simple and accurate method could be employed in field virus testing.

We report that the mammalian 5-methylcytosine (5mC) oxidase Tet3 exists as three major isoforms and characterized the full-length isoform containing an N-terminal CXXC domain (Tet3FL). This CXXC domain binds to unmethylated CpGs, but, unexpectedly, its highest affinity is toward 5-carboxylcytosine (5caC). We determined the crystal structure of the CXXC domain-5caC-DNA complex, revealing the structural basis of the binding specificity of this domain as a reader of CcaCG sequences. Mapping of Tet3FL in neuronal cells shows that Tet3FL is localized precisely at the transcription start sites (TSSs) of genes involved in lysosome function, mRNA processing, and key genes of the base excision repair pathway. Therefore, Tet3FL may function as a regulator of 5caC removal by base excision repair. Active removal of accumulating 5mC from the TSSs of genes coding for lysosomal proteins by Tet3FL in postmitotic neurons of the brain may be important for preventing neurodegenerative diseases.

Although the motivating role of feedback and progress indicators is understood in formal learning, their role in supporting incidental mobile learning is less well understood. In this paper we argue that well-designed feedback and progress indicators (FPIs) offer guidance and structure that may encourage mobile app users to move from fragmented learning episodes towards a longer term, reflective learning journey. Drawing from relevant literature we consider how FPIs can be used in the EU-funded MASELTOV project which explores how a suite of smartphone apps can support recent immigrants to Europe to become integrated in their new cities. These apps allow learning episodes to be part of daily activities and interactions. The authors discuss what kinds of FPIs should be provided and introduce the SCAMP model which emphasises five types of FPIs--Social, Cognitive, Affective, Motivational and Progress. Finally, the authors provide examples of FPIs that will be used in the MASELTOV project.