Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Disentangling hexaploid genetics : towards DNA-informed breeding for postharvest performance in chrysanthemum
    Geest, Geert van - \ 2017
    Wageningen University. Promotor(en): R.G.F. Visser, co-promotor(en): U. van Meeteren; P.F.P. Arens. - Wageningen : Wageningen University - ISBN 9789463436427 - 142
    chrysanthemum - plant breeding - postharvest quality - hexaploidy - polyploidy - quantitative trait loci - phenotypes - linkage mapping - metabolomics - polymorphism - dna - chrysanthemum - plantenveredeling - kwaliteit na de oogst - hexaploïdie - polyploïdie - loci voor kwantitatief kenmerk - fenotypen - koppelingskartering - metabolomica - polymorfisme - dna

    DNA-informed selection can strongly improve the process of plant breeding. It requires the detection of DNA polymorphisms, calculation of genetic linkage, access to reliable phenotypes and methods to detect genetic loci associated with phenotypic traits of interest. Cultivated chrysanthemum is an outcrossing hexaploid with an unknown mode of inheritance. This complicates the development of resources and methods that enable the detection of trait loci. Postharvest performance is an essential trait in chrysanthemum, but is difficult to measure. This makes it an interesting but challenging trait to phenotype and detect associated genetic loci. In this thesis I describe the development of resources and methods to enable phenotyping for postharvest performance, genetic linkage map construction and detection of quantitative trait loci in hexaploid chrysanthemum.

    Postharvest performance is a complicated trait because it is related to many different disorders that reduce quality. One of these disorders in chrysanthemum is disk floret degreening, which occurs after long storage. In chapter 2, we show that degreening can be prevented by feeding the flower heads with sucrose, suggesting carbohydrate starvation plays a role in the degreening process. To investigate the response to carbohydrate starvation of genotypes with different sensitivity to disk floret degreening, we investigated the metabolome of sugar-fed and carbohydrate-starved disk florets by 1H-NMR and HPAEC. We show that the metabolome is severely altered at carbohydrate starvation. In general, starvation results in an upregulation of amino acid and secondary metabolism. Underlying causes of genotypic differences explaining variation in disk floret degreening in the three investigated genotypes remained to be elucidated, but roles of regulation of respiration rate and camphor metabolism were posed as possible candidates.

    In chapter 3, disk floret degreening was found to be the most important postharvest disorder after 3 weeks of storage among 44 white chrysanthemum cultivars. To investigate the inheritance of disk floret degreening, we crossed two genotypes with opposite phenotypic values of both disk floret degreening and carbohydrate content to obtain a population segregating for disk floret degreening. To phenotype the cultivar panel and the bi-parental population precisely and in a high throughput manner, we developed a method that quantified colour of detached capitula over time. This method was validated with visual observations of disk floret degreening during vase life tests. In a subset of the bi-parental population we measured carbohydrate content of the disk florets at harvest. The amount of total carbohydrates co-segregated with sensitivity to degreening, which shows that the difference in disk floret degreening sensitivity between the parents could be explained by their difference in carbohydrate content. However, the correlation was rather weak, indicating carbohydrate content is not the only factor playing a role.

    In order to develop resources for DNA-informed breeding, one needs to be able to characterize DNA polymorphisms. In chapter 4, we describe the development of a genotyping array containing 183,000 single nucleotide polymorphisms (SNPs). These SNPs were acquired by sequencing the transcriptome of 13 chrysanthemum cultivars. By comparing the genomic dosage based on the SNP assay and the dosage as estimated by the read depth from the transcriptome sequencing data, we show that alleles are expressed conform the genomic dosage, which contradicts to what is often found in disomic polyploids. In line with this finding, we conclusively show that cultivated chrysanthemum exhibits genome-wide hexasomic inheritance, based on the segregation ratios of large numbers of different types of markers in two different populations.

    Tools for genetic analysis in diploids are widely available, but these have limited use for polyploids. In chapter 5, we present a modular software package that enables genetic linkage map construction in tetraploids and hexaploids. Because of the modularity, functionality for other ploidy levels can be easily added. The software is written in the programming language R and we named it polymapR. It can generate genetic linkage maps from marker dosage scores in an F1 population, while taking the following steps: data inspection and filtering, linkage analysis, linkage group assignment and marker ordering. It is the first software package that can handle polysomic hexaploid and partial polysomic tetraploid data, and has advantages over other polyploid mapping software because of its scalability and cross-platform applicability.

    With the marker dosage scores of the bi-parental F1 population from the genotyping array and the developed methods to perform linkage analysis we constructed an integrated genetic linkage map for the hexaploid bi-parental population described in chapter 3 and 4. We describe this process in chapter 6. With this integrated linkage map, we reconstructed the inheritance of parental haplotypes for each individual, and expressed this as identity-by-descent (IBD) probabilities. The phenotypic data on disk floret degreening sensitivity that was acquired as described in chapter 3, was used in addition to three other traits to detect quantitative trait loci (QTL). These QTL were detected based on the IBD probabilities of 1 centiMorgan intervals of each parental homologue. This enabled us to study genetic architecture by estimating the effects of each separate allele within a QTL on the trait. We showed that for many QTL the trait was affected by more than two alleles.

    In chapter 7, the findings in this thesis are discussed in the context of breeding for heterogeneous traits, the implications of the mode of inheritance for breeding and the advantages and disadvantages of polyploidy in crop breeding. In conclusion, this thesis provides in general a significant step for DNA-informed breeding in polysomic hexaploids, and for postharvest performance in chrysanthemum in particular.

    How virtual shade sheds light on plant plasticity
    Bongers, Franca J. - \ 2017
    Wageningen University. Promotor(en): N.P.R. Anten, co-promotor(en): R. Pierik; J.B. Evers. - Wageningen : Wageningen University - ISBN 9789463432047 - 140
    planten - fenotypen - fenotypische variatie - modellen - arabidopsis - natuurlijke selectie - schaduw - reacties - concurrentie tussen planten - licht - plants - phenotypes - phenotypic variation - models - arabidopsis - natural selection - shade - responses - plant competition - light

    Phenotypic plasticity is the ability of a genotype to express multiple phenotypes in accordance with different environments. Although variation in plasticity has been observed, there is limited knowledge on how this variation results from natural selection. This thesis analyses how variation in the level of plasticity influences light competition between plants and how this variation could result from selection, driven by light competition, in various environments. As an exemplary case of phenotypic plasticity, this thesis focusses on phenotypic responses of the annual rosette plant Arabidopsis thaliana (Brassicaceae) in response to the proximity of neighbour plants, as signalled through the red : far—red (R:FR) ratio, which are responses associated with the shade avoidance syndrome (SAS).

    Plant experiments were conducted to measure variation in these plastic responses and a functional-structural plant (FSP) model was created that simulates plant structures in 3D and includes these organ-level plastic responses while simulating explicitly a heterogeneous light environment. Simulating individual plants that explicitly compete for light, while their phenotype changes through plasticity, gave insights in the role of the level of phenotypic plasticity and site of signal perception on plant competitiveness. In addition, an analysis on how natural selection in different environments acts on the level of plasticity was performed by combining FSP simulations and evolutionary game theoretical (EGT) principles.

    Differences in transcriptional responses to acute and chronic dietary interventions with fatty acids
    Matualatupauw, Juri C. - \ 2017
    Wageningen University. Promotor(en): A.H. Kersten, co-promotor(en): L.A. Afman; J. Bouwman. - Wageningen : Wageningen University - ISBN 9789463432078 - 172
    fatty acids - gene expression - genotyping - phenotypes - nutritional intervention - transcriptomics - fish oils - apolipoprotein e - adipose tissue - microarrays - polymerase chain reaction - vetzuren - genexpressie - genotyping - fenotypen - maatregel op voedingsgebied - transcriptomica - visoliën - apolipoproteïne e - vetweefsel - microarrays - polymerase-kettingreactie

    Various types of dietary fatty acids have different effects on human health. The aim of this thesis was to increase our understanding of the molecular mechanisms underlying the effects of dietary fatty acids. To do this, we examined changes in whole genome gene expression profiles upon both acute as well as longer term dietary fatty acid interventions. Furthermore, from previous research, it is clear that large inter-individual differences in the response to dietary fatty acids exist. We used whole genome gene expression analyses to increase our understanding of the mechanisms underlying some of these inter-individual differences.

    Many modifiable and non-modifiable factors can be the cause of these inter-individual differences. In chapter 2, we reviewed all studies that examined differences in the transcriptional response to dietary interventions based on the presence of one of these factors. These include gender, age, BMI, body composition, blood lipid levels and gut microbial composition. We conclude that transcriptome analyses are well-suited for studying the underlying mechanisms behind these differences in the response to diet. Nevertheless, the number of studies that use this approach remains limited.

    Another factor that may modify the response to a dietary intervention is genetics, e.g. the apolipoprotein E4 (APOE4) variant. People who carry the APOE4 allele have an increased risk of cardiovascular disease. Fish-oil supplementation may help in the prevention of cardiovascular disease, though inter-individual differences in the response to n-3 polyunsaturated fatty acids on gene expression profiles have been observed. In chapter 3, we aimed to assess the impact of APOE4 on peripheral blood mononuclear cell (PBMC) whole genome gene expression at baseline and following a 6-month fish-oil intervention. We observed increased gene expression of IFN signaling and cholesterol biosynthesis pathways in APOE4 carriers, which might explain part of the association between APOE4 and CVD. Furthermore, fish-oil supplementation may be beneficial by decreasing interferon signalling-related gene expression in APOE4 carriers.

    Another long-term dietary intervention with fatty acids was studied in chapter 4. We examined the effect of a 12-week high medium-chain saturated fatty acid diet on subcutaneous adipose tissue gene expression profiles. We observed increased expression of genes involved in oxidative energy metabolism and decreased inflammation-related gene expression due to the high medium-chain saturated fatty acid intake. Considering the role of the adipose tissue in sustaining the low-grade inflammation that is associated with obesity, these findings may be indicative of a more anti-inflammatory phenotype of the adipose tissue. We concluded that medium-chain saturated fatty acids may potentially have beneficial effects on adipose tissue functioning.

    Besides studying the effects of long-term interventions with fatty acids on whole genome gene expression, we also examined the effects of acute high-fat challenges. In chapter 5, we determined the additional value of determining whole genome gene expression changes in response to a high-fat challenge compared to assessment at fasting only. In addition, we aimed to identify whether a 4 week high-fat high-calorie diet can induce a shift in gene expression profiles in healthy subjects towards a metabolic syndrome-like gene expression profile. We found that fasting whole blood whole genome gene expression profiles are highly responsive to a 4-week high-fat high-calorie diet, with changes in in the direction of a metabolic syndrome-like gene expression profile. High-fat challenge responses in healthy subjects show only minimal changes in gene expression upon the dietary intervention and a marginal shift in the direction of the metabolic syndrome. We concluded that fasting gene expression profiles are more responsive compared to high-fat challenge responses to a 4-week high-fat high-calorie diet.

    Besides chapter 5, several other studies have also examined changes in whole genome gene expression in blood cells induced by high-fat challenges. In chapter 6, we combined microarray data from four high-fat challenge studies varying in study population, challenge composition and research laboratory. By performing this meta-analysis, we showed a general PBMC whole genome gene expression response to a high-fat challenge. We concluded that a meta-analysis provides added value for the discovery of consistently differentially expressed genes and pathways compared to selecting only those genes and pathways that are identified in all separate studies.

    In conclusion, in this thesis we showed differences in the whole genome gene expression response to fish-oil supplementation in PBMCs of APOE4 carriers vs non-carriers. Furthermore, the effects on whole genome gene expression of the two long-term dietary interventions, i.e. the fish-oil supplementation in PBMCs of APOE4 carriers and the high medium-chain saturated fatty acid diet in adipose tissue, may be beneficial by downregulation of gene expression related to inflammation. We also showed that whole genome gene expression responses to high-fat challenges are affected by a 4-week high-fat high-calorie diet, though changes in fasting gene expression profiles are much more pronounced. Finally, we showed the value of meta-analysis of microarray data in high-fat challenge studies for identifying the general response to a high-fat challenge.

    Using probabilistic graphical models to reconstruct biological networks and linkage maps
    Wang, Huange - \ 2017
    Wageningen University. Promotor(en): F.A. Eeuwijk, co-promotor(en): J. Jansen. - Wageningen : Wageningen University - ISBN 9789463431538 - 150
    probabilistic models - models - networks - linkage - mathematics - statistics - quantitative trait loci - phenotypes - simulation - waarschijnlijkheidsmodellen - modellen - netwerken - koppeling - wiskunde - statistiek - loci voor kwantitatief kenmerk - fenotypen - simulatie

    Probabilistic graphical models (PGMs) offer a conceptual architecture where biological and mathematical objects can be expressed with a common, intuitive formalism. This facilitates the joint development of statistical and computational tools for quantitative analysis of biological data. Over the last few decades, procedures based on well-understood principles for constructing PGMs from observational and experimental data have been studied extensively, and they thus form a model-based methodology for analysis and discovery. In this thesis, we further explore the potential of this methodology in systems biology and quantitative genetics, and illustrate the capabilities of our proposed approaches by several applications to both real and simulated omics data.

    In quantitative genetics, we partition phenotypic variation into heritable, genetic, and non-heritable, environmental, parts. In molecular genetics, we identify chromosomal regions that drive genetic variation: quantitative trait loci (QTLs). In systems genetics, we would like to answer the question of whether relations between multiple phenotypic traits can be organized within wholly or partially directed network structures. Directed edges in those networks can be interpreted as causal relationships, causality meaning that the consequences of interventions are predictable: phenotypic interventions in upstream traits, i.e. traits occurring early in causal chains, will produce changes in downstream traits. The effect of a QTL allele can be considered to represent a genetic intervention on the phenotypic network. Various methods have been proposed for statistical reconstruction of causal phenotypic networks exploiting previously identified QTLs. In chapter 2, we present a novel heuristic search algorithm, namely the QTL+phenotype supervised orientation (QPSO) algorithm, to infer causal relationships between phenotypic traits. Our algorithm shows good performance in the common, but so far uncovered case, where some traits come without QTLs. Therefore, our algorithm is especially attractive for applications involving expensive phenotypes, like metabolites, where relatively few genotypes can be measured and population size is limited.

    Standard QTL mapping typically models phenotypic variations observable in nature in relation to genetic variation in gene expression, regardless of multiple intermediate-level biological variations. In chapter 3, we present an approach integrating Gaussian graphical modeling (GGM) and causal inference for simultaneous modeling of multilevel biological responses to DNA variations. More specifically, for ripe tomato fruits, the dependencies of 24 sensory traits on 29 metabolites and the dependencies of all the sensory and metabolic traits further on 21 QTLs were investigated by three GGM approaches including: (i) lasso-based neighborhood selection in combination with a stability approach to regularization selection, (ii) the PC-skeleton algorithm and (iii) the Lasso in combination with stability selection, and then followed by the QPSO algorithm. The inferred dependency network which, though not essentially representing biological pathways, suggests how the effects of allele substitutions propagate through multilevel phenotypes. Such simultaneous study of the underlying genetic architecture and multifactorial interactions is expected to enhance the prediction and manipulation of complex traits. And it is applicable to a range of population structures, including offspring populations from crosses between inbred parents and outbred parents, association panels and natural populations.

    In chapter 4, we report a novel method for linkage map construction using probabilistic graphical models. It has been shown that linkage map construction can be hampered by the presence of genotyping errors and chromosomal rearrangements such as inversions and translocations. Our proposed method is proven, both theoretically and practically, to be effective in filtering out markers that contain genotyping errors. In particular, it carries out marker filtering and ordering simultaneously, and is therefore superior to the standard post-hoc filtering using nearest-neighbour stress. Furthermore, we demonstrate empirically that the proposed method offers a promising solution to genetic map construction in the case of a reciprocal translocation.

    In the domain of PGMs, Bayesian networks (BNs) have proven, both theoretically and practically, to be a promising tool for the reconstruction of causal networks. In particular, the PC algorithm and the Metropolis-Hastings algorithm, which are representatives of mainstream methods to BN structure learning, are reported to have been successfully applied to the field of biology. In view of the fact that most biological systems exist in the form of random network or scale-free network, in chapter 5 we compare the performance of the two algorithms in constructing both random and scale-free BNs. Our simulation study shows that for either type of BN, the PC algorithm is superior to the M-H algorithm in terms of timeliness; the M-H algorithm is preferable to the PC algorithm when the completeness of reconstruction is emphasized; but when the fidelity of reconstruction is taken into account, the better one of the two algorithms varies from case to case. Moreover, whichever algorithm is adopted, larger sample sizes generally permit more accurate reconstructions, especially in regard to the completeness of the resulting networks.

    Finally, chapter 6 presents a further elaboration and discussion of the key concepts and results involved in this thesis.

    The fatter the better : selecting microalgae cells for outdoor lipid production
    Dominguez Teles, I. - \ 2016
    Wageningen University. Promotor(en): Rene Wijffels, co-promotor(en): Maria Barbosa; Dorinde Kleinegris. - Wageningen : Wageningen University - ISBN 9789462578821 - 164
    algae - chlorococcum - lipids - lipogenesis - fat - production - phenotypes - inoculum - diameter - cells - sorting - algen - chlorococcum - lipiden - lipogenese - vet - productie - fenotypen - entstof - diameter - cellen - sorteren

    In chapter 1 we introduce microalgae, photosynthetic microorganisms with potential to replace commodities (such as food, feed, chemicals and fuels). Production costs are still high, reason why microalgae are still only economically feasible for niche markets. We suggest to borrow the concept of plant domestication to select industrial microalgae cells. Two approaches can be successfully used to domesticate microalgae: adaptive laboratory evolution (ALE) and fluorescence assisted cell sorting (FACS). ALE takes advantage of the natural adaptability of microorganisms to different environments, while FACS actually select cells with specific phenotypes. This thesis aimed to select cells of Chlorococcum littorale with improved phenotypes, assuming that these cells could establish new populations with increased industrial performance.

    In Chapter 2 we wanted to know what happened during time to biomass and lipid productivities of Chlorococcum littorale repeatedly subjected to N-starvation. We tested 2 different cycles of N-starvation, short (6 days) and long (12 days). Short cycles didn’t affect lipid productivity, highlighting the potential of C. littorale to be produced in semi-continuous cultivation. Repeated cycles of N-starvation could have caused adaptations of the strain. Hence, we also discussed the implications of using repeated N-starvation for adaptive laboratory evolution (ALE) experiments. Chapter 3 introduces a method to detect and to select microalgae cells with increased lipid content. The method requires only the fluorescence dye Bodipy505/515 dissolved in ethanol, and the method was designed to maintain cellular viability so the cells could be used to produce new inoculum. In chapter 4 we evaluated a question that emerged while deciding which criteria to use to sort lipid-rich cells: does cellular size affects lipid productivity of C. littorale? We hypothesized that cells with different diameters have different division rates, which could affect lipid productivity. Therefore, we assessed the influence of cell diameter, as a sorting parameter, on both biomass and lipid productivity of Chlorococcum littorale (comparing populations before and after sorting, based on different diameters). Results showed that the size of vegetative cells doesn’t affect the lipid productivity of C. littorale. In chapter 5 we present a strategy to sort cells of C. littorale with increased TAG productivity using the method developed at chapter 3. Both the original and the sorted population with the highest lipid productivity (namely, S5) were compared under simulated Dutch summer conditions. The results confirmed our data from experiments done under continuous light: S5 showed a double TAG productivity. Our results showed also that the selected phenotype was stable (1.5 year after sorting) and with potential to be used under industrial conditions. In chapter 6 we extrapolated our results (indoor and outdoor) to other climate conditions. We ran simulations changing the light conditions to four different locations worldwide (the Netherlands, Norway, Brazil and Spain) to estimate both biomass and TAG productivities. Results indicated that biomass yields were reduced at locations with higher light intensities (Brazil/Spain) when compared with locations with lower light intensities (Norway/Netherlands). Hence, the choice of location should not be based on light intensity, but on how stable irradiation is. Chapter 7 is the general discussion of the thesis, demonstrating that both ALE and FACS are effective approaches to select industrial microalgae cells. We also present our view on how ALE and FACS could further improve microalgae strains for industry.

    Plant responses to multiple herbivory : phenotypic changes and their ecological consequences
    Li, Yehua - \ 2016
    Wageningen University. Promotor(en): Marcel Dicke, co-promotor(en): Rieta Gols. - Wageningen : Wageningen University - ISBN 9789462578043 - 165
    brassica oleracea - brevicoryne brassicae - aphidoidea - caterpillars - insect pests - pest resistance - defence mechanisms - phenotypes - insect plant relations - parasitoids - natural enemies - herbivore induced plant volatiles - plant-herbivore interactions - genetic variation - brassica oleracea - brevicoryne brassicae - aphidoidea - rupsen - insectenplagen - plaagresistentie - verdedigingsmechanismen - fenotypen - insect-plant relaties - parasitoïden - natuurlijke vijanden - herbivoor-geinduceerde plantengeuren - plant-herbivoor relaties - genetische variatie

    This thesis explores whether aphid-infestation interferes with the plant response to chewing herbivores and whether this impacts performance and behaviour of individual chewing insect herbivores and their natural enemies, as well as the entire insect community. I investigated this using three wild cabbage populations (Brassica oleracea) that are known to differ in inducible secondary chemistry, to reveal whether patterns were consistent.

    A literature review on recent developments in the field of plant interactions with multiple herbivores (Chapter 2) addressed how plant traits mediate interactions with various species of the associated insect community and their dynamics. In addition, the mechanisms underlying phenotypic changes in response to different herbivores were discussed from the expression of defence-related genes, phytohormones and secondary metabolites in plants to their effects on the performance and behaviour of individual insects as well as the entire insect community. In Chapter 3, I investigated the effects of early-season infestation by the aphid Brevicoryne brassicae on the composition and dynamics of the entire insect community throughout the season in a garden experiment replicated in two consecutive years. Aphid infestation in the early season only affected a subset of the community, i.e. the natural enemies of aphids, but not the chewing herbivores and their natural enemies. Moreover, the effects were only significant in the first half (June & July), but waned in the second half of the season (August & September). The effect of aphid infestation on the community of natural enemies also varied among the cabbage populations. Chapter 4 investigated the effects of aphid infestation on plant direct defences against chewing herbivores in laboratory experiments by comparing the performance of chewing herbivores and their parasitoids on aphid-infested and aphid-free plants. The performance of the specialist herbivore Plutella xylostella and its parasitoid Diadegma semiclausum was better on plants infested with aphids than on aphid-free plants, whereas the performance of the generalist herbivore Mamestra brassicae and its parasitoid Microplitis mediator was not affected by aphid infestation. These results suggest that aphid induced changes in plant traits may differentially affect the performance of leaf-chewing herbivore species attacking the same host plant, and also varied among the cabbage populations. Chapter 5 examined the effects of B. brassicae aphid infestation on plant indirect defences against chewing herbivores. In a two-choice olfactometer bioassay, preference behaviour for volatiles emitted by plants infested with hosts alone and those emitted by plants infested with aphids and hosts was compared for D. semiclausum and M mediator, larval endoparasitoids of caterpillars of P. xylostella and M. brassicae, respectively. In addition, the headspace volatiles emitted by host-infested and dually-infested plants were collected and analyzed. Co-infestation with aphids differentially affected volatile-mediated foraging behaviour of the two parasitoid species in an infestation period-dependent manner. Diadegma semiclausum preferred dually infested plants over host-infested plants when aphids infested the plants for a short time period, i.e. 7 days, but the volatile preference of D. semiclausum was reversed when aphid infestation was extended to 14 days. In contrast, M. mediator consistently preferred volatiles emitted by the dually-infested plants over those emitted by host-infested plants. The patterns of preference behaviour of the two wasp species were consistent across the three cabbage populations. Interestingly, the emission rate of most volatile compounds was reduced in plants dually-infested with caterpillars and aphids compared to singly-infested with caterpillars. This study showed that aphid infestation increased plant indirect defences against caterpillars, but depended on the aphid infestation period and specific caterpillar-parasitoid association. We hypothesized a negative interference of aphid infestation on plant defences against chewing herbivores based on previously reported SA-JA antagonism. In Chapter 6, we assessed the activation of SA and JA signaling pathways in plants infested by both aphids (B. brassicae) and various caterpillar species (P. xylostella, M. brassicae and Pieris brassicae) in different time sequences by quantifying transcription levels of the SA- and JA-responsive marker genes, PR-1 and LOX respectively. The results did not provide support for SA-JA antagonism. Compared to single infestation with each of the herbivore species, dual infestation with aphid and caterpillars had no interactive effects on the transcription levels of the SA- and JA-responsive maker genes, regardless of the temporal sequence of aphid and caterpillar attack, or the identity of the attacking caterpillar species.

    The findings of this thesis contribute to our understanding of plant responses to herbivory by insect species belonging to different feeding guilds and their ecological effects on other associated community members. Aphid infestation may interfere with plant direct and indirect defences against leaf-chewing herbivores at the individual species level, but the effects are species-specific and also depend on the infestation period of aphids. Early-season aphid infestation may further affect the composition of the insect community, but the effect is smaller influencing only a subset of the community compared to early infestation by chewing herbivores. The molecular mechanism underlying plant responses to both phloem-feeding and leaf-chewing herbivores are complex and require the investigation of a range of genes involved in JA- and SA-mediated defence signal transduction. Plant interact with multiple herbivores at different levels of biological organization ranging from the subcellular level to the individual and the community level, and an integrated multidisciplinary approach is required to investigate plant-insect interactions.

    Natural genetic variation in Arabidopsis thaliana photosynthesis
    Flood, P.J. - \ 2015
    Wageningen University. Promotor(en): Maarten Koornneef, co-promotor(en): Mark Aarts; Jeremy Harbinson. - Wageningen : Wageningen University - ISBN 9789462575004 - 278
    arabidopsis thaliana - genetische variatie - fotosynthese - genomen - chlorofyl - fenotypen - arabidopsis thaliana - genetic variation - photosynthesis - genomes - chlorophyll - phenotypes

    Oxygenic photosynthesis is the gateway of the sun’s energy into the biosphere, it is where light becomes life. Genetic variation is the fuel of evolution, without it natural selection is powerless and adaptation impossible. In this thesis I have set out to study a relatively unexplored field which sits at the intersection of these two topics, namely natural genetic variation in plant photosynthesis. To begin I reviewed the available literature (Chapter 2), from this it became clear that the main bottleneck restricting progress was the lack of high-throughput phenotyping platforms for photosynthesis. To address this an automated high-throughput chlorophyll fluorescence phenotyping system was developed, which could measure 1440 plants in less than an hour for ΦPSII, a measure of photosynthetic efficiency (Chapter 3). Using this phenotyping platform I screened five populations of Arabidopsis thaliana. Three of these populations resulted from bi-parental crosses and segregated for only two genomes, using these I conducted family mapping (Chapter 4). The final two populations were composed of natural, field collected, accessions and were analysed using a genome wide association approach (Chapter 5). The family mapping approach had greater statistical power due to within population replication and the genome wide association approach had higher mapping resolution due to historical recombination. Both approaches were used to identify genomic regions (loci) which were responsible for some of the variation in photosynthesis observed. The number and average effect of these loci was used to infer the genetic architecture of photosynthesis as a highly complex polygenic trait for which there are many loci of very small effect. In addition to screening these large populations a smaller subset of 18 lines was assayed for natural variation in phosphorylation of photosystem II (PSII) proteins in response to changing light (Chapter 6). This exploratory study indicated that this process shows considerable variation and may be important for adaptation of the photosynthetic apparatus to photosynthetic extremes. The genetic mapping studies just described, focus exclusively on genetic variation in the nuclear genome, whilst this contains the majority of the plants genetic information there is also a store of genetic information in the chloroplast and mitochondria. These genetic repositories contain genes which are essential for photosynthesis and energy metabolism. Any variation in these genes could have a large impact on photosynthesis. To study natural variation in these genomes I developed a new population of reciprocal nuclear-organellar hybrids (cybrids) which could be used to study the effect of genetic variation in organelles whilst controlling for nuclear genetic variation (Chapter 7). Preliminary results indicate that this resource will be of great use in disentangling natural genetic variation in nucleo-organelle interactions. Finally I looked at one chloroplast encoded photosynthetic mutation in more detail (Chapter 8). This mutation had evolved in response to herbicide application and had spread along British railways. When studying this population of resistant plants I found empirical evidence for organelle mediated nuclear genetic hitchhiking. This is a previously undescribed evolutionary phenomenon and is likely to be quite common. In conclusion there is an abundance of genetic variation in photosynthesis which can be used to improve the trait for agriculture and provide insights into novel evolutionary phenomena in the field.

    Role of anti-competitor toxins in the origin and maintenance of diversity in Saccharomyces yeast microbial populations
    Pieczynska, M.D. - \ 2015
    Wageningen University. Promotor(en): Bas Zwaan, co-promotor(en): Arjan de Visser. - Wageningen : Wageningen University - ISBN 9789462573093 - 123
    gisten - rna-virussen - toxinen - symbiose - toxiciteit - co-evolutie - fenotypen - fermentatie - yeasts - rna viruses - toxins - symbiosis - toxicity - coevolution - phenotypes - fermentation


    Saccharomyces cells occasionally carry cytoplasmic ds-RNA “killer” viruses coding for low-mass proteins, which upon secretion to the environment can kill related cells that do not carry the viral particles. Such killer viruses are not infectious, and can spread only through cell division and during mating. Three principal classes of Saccharomyces viruses (ScV-M1, ScV-M2 and ScV-M28) belonging to the Totiviridae family have been characterised, each capable of forming a specific anti-competitor toxin and corresponding antidote. Presumably, toxic killing provides competitive benefits to the yeast host. However, the ecological and evolutionary significance of toxin production remains poorly understood. For example, it is unknown where yeast killers occur and at what frequency, how evolvable killing ability is, whether it is constrained by possible trade-offs with resource competitive ability and how it is shaped by interactions with toxin-sensitive competitors. Also unknown is how stable yeast-virus symbioses are, and how coevolution between host and virus may affect this stability and the killing phenotype itself. It is believed that killer yeasts are common based on the fact that they have been found among yeasts isolated from different sources over several decades. In chapter 2, we assay two large yeast collections from diverse habitats, including nature and man-made habitats (in total 136 strains with known genome sequences), for killer phenotype and toxin resistance. We find that ~10.3% carry a killer virus, while about 25% are resistant to at least one of the three known killer toxins (12.5% to different combinations of two and ~9% to all three), most likely due to chromosomal mutations. Analyses of their evolutionary relationship indicate that host-virus associations are relatively short lived, whereas the relatively high frequency of resistance suggests that toxins have a substantial impact on yeast evolution.

    In order to understand the ecological and evolutionary role of toxin production, it is essential to reliably assess the killing rate of toxin producers by measuring how many toxin-sensitive individuals are killed by a single toxin producer during a given time interval. To identify a convenient method with high sensitivity and reproducibility, in chapter 3 we perform a systematic comparative analysis of four methods, including the conventional “Halo method” and three more quantitative liquid assays. We apply these methods to a set of three known yeast killer strains (K1, K2 and K28) and find that the easy applicable Halo method provides the most sensitive and reproducible killing rate estimates (with best discrimination between killer strains).

    Understanding the evolution of the yeast-virus association is crucial for a full understanding of the ecological and evolutionary role of killer strains. In chapter 4, we present experimental tests of the strength of the dependence of yeast host strains on their killer viruses. We cross-infect several viruses among killer strains of the genus Saccharomyces – all expressing the K1-type toxin, and test native and new combinations for the strength of host-virus co-adaptation. We find explicit host-virus co-adaptation, because native yeasts hosts display the highest toxicity and highest stability of killer viruses relative to hosts carrying non-native viruses. Even stronger, we find that curing these wild killer yeasts from their virus reduces their competitive fitness, despite initial fitness costs of viral carriage reported for constructed killer strains. These results demonstrate co-adaptation of host and virus in the natural killer strains resulting in their dependence on the killer virus. To explore the evolutionary costs and benefits of virus carriage and toxin production, and understand whether they are shaped by the coevolution between host and virus and the presence of toxin-sensitive competitors in the environment, we conduct a series of laboratory experiments where we manipulate the opportunity for coevolution (chapter 5). Analyses of killing ability, toxin sensitivity and fitness (i.e. resource competitive ability), show rapid reciprocal changes in killer and sensitive strain when coevolution is allowed, modulated by the rapid invasion of toxin-resistant mutants and subsequent reduction of killing ability. Remarkably, we find that the rapid invasion of toxin-resistant mutants involves two mutational steps, the first being a mutation showing a meiotic drive phenotype as well as a strong fitness benefit in heterozygotes, the second the resistance mutation. Shifts in the competitive fitness of evolved killer isolates with increased killing ability show a clear trade-off between killing rate and resource competitive ability, indicating that resource and interference competitive ability are alternative competitive strategies. Moreover, by cross-infecting the killer virus between the ancestral and an evolved strain, we are able to demonstrate the rapid co-adaptation between host and killer virus, supporting our previous findings of co-adaptive responses in wild yeast killers (chapter 4).

    Our analyses are based on screens of natural isolates, laboratory evolution experiments and phenotypic analyses, complemented by classical genetics. To more fully understand the reciprocal nature and molecular mechanisms of adaptive responses, genome analyses are required. The motivation for such analyses and other follow-up studies are proposed in chapter 6. My studies show the usefulness of the killer yeast system to address questions related to interference competition and coevolution, which may proof valuable also given potential applications of killer yeasts in the fermentation industry.

    Registreren: meten is weten!
    Maurice - Van Eijndhoven, M.H.T. ; Oldenbroek, J.K. - \ 2015
    Zeldzaam huisdier 40 (2015)2. - ISSN 0929-905X - p. 10 - 11.
    rassen (dieren) - dierveredeling - selectie - registratie - selectief fokken - fenotypen - fokdoelen - stamboeken - databanken - breeds - animal breeding - selection - registration - selective breeding - phenotypes - breeding aims - herdbooks - databases
    Om te kunnen selecteren op bepaalde kenmerken moeten deze ‘meetbaar’
    zijn en worden geregistreerd. Pas dan wordt zichtbaar of er echte
    verbeteringen richting fokdoel worden bereikt in volgende generaties.
    In dit tweede artikel lichten we toe waarom het belangrijk is om goed
    te registeren en hoe een gedegen registratiesysteem eruitziet.
    Feather pecking and monoamines - a behavioral and neurobiological approach
    Kops, M.S. - \ 2014
    Wageningen University; Utrecht University. Promotor(en): B. Olivier; O. Güntürkün, co-promotor(en): S.M. Korte; Liesbeth Bolhuis. - Utrecht, The Netherlands : Utrecht University - ISBN 9789039361283 - 172
    pluimveehouderij - hennen - verenpikken - diergedrag - dierenwelzijn - pluimvee - diergezondheid - dierlijke productie - serotonine - dopamine - fenotypen - genotypen - neurotransmitters - invloeden - poultry farming - hens - feather pecking - animal behaviour - animal welfare - poultry - animal health - animal production - serotonin - dopamine - phenotypes - genotypes - neurotransmitters - influences
    Severe feather pecking (SFP) remains one of the major welfare issues in laying hens. SFP is the pecking at and pulling out of feathers, inflicting damage to the plumage and skin of the recipient. The neurobiological profile determining the vulnerability of individual hens to develop into a severe feather pecker is unknown, although brain monoamines such as serotonin (5-HT) and dopamine (DA) seem to play a role. Previous studies related lower 5-HT and DA turnover ratios to an increased risk to develop SFP.In this thesis, monoamine levels in brain areas involved in emotional regulation and motor control were compared between phenotypically and genetically selected high and low feather peckers at different ages. It was found that adult high feather peckers had higher monoaminergic activity (lower metabolite levels and/or turnover ratios) in comparison to low feather peckers, which is in contrast with results on young hens. Differences were seen in several brain areas, namely the dorsal thalamus, medial striatum, amygdala, caudocentral nidopallium, and the somatomotor arcopallium, but to a lesser extend or not in the caudolateral nidopallium and the hippocampus. To investigate the exact neurobiological mechanism behind severe feather pecking further extracellular levels of 5-HT and DA and their metabolites were measured by in vivo microdialysis. Up till now, microdialysis has only been executed in young chickens, but this thesis describes the first microdialysis study performed in adult laying hens. It was found that adult severe feather peckers had a higher baseline release of 5-HT in the caudal nidopallium, a large associative area in the chicken’s forebrain. This result could not be explained by the amount of 5-HT presynaptically stored, as both high and low SFP lines displayed a similar 5-HT release after d-fenfluramine administration. This confirms that genetic selection on SFP has altered the serotonergic system in feather pecking-phenotypes. With clear phenotypic and genotypic differences in brain areas related to emotional regulation and motor control, it can be assumed that brain deficits at a young age increase an individual’s vulnerability to stressful environmental changes, which is associated with the prevalence of SFP later in life. The cause of the inversion of neurochemical patterns in young and adult high and low feather pecking hens remains to be elucidated. Perhaps this inversion is caused by development itself. On the other hand, higher behavioral patterns (SFP and other types of allopecking) observed in the high feather pecking chickens might have influenced the monoaminergic activity since the brain influences behavior and vice versa. Altogether, this thesis demonstrates the importance of considering the impact of genetic selection and also environmental conditions on brain neurotransmission and with that, on the vulnerability of individual chickens to develop SFP. Both the serotonergic and dopaminergic systems are involved in the development of SFP. With SFP being a multifactorial problem both genotype and phenotype have to be taken into account. Furthermore, in vivo microdialysis is a valuable approach to investigate why individual laying hens start SFP. This will lead to further understanding and ultimately in the reduction of SFP.
    Potato genetical genomics: investigating the genetic basis of primary metabolism and its relationship to the phenotype
    Carreño Quintero, N. - \ 2013
    Wageningen University. Promotor(en): Harro Bouwmeester; Richard Visser, co-promotor(en): Joost Keurentjes; Christian Bachem. - Wageningen : Wageningen University - ISBN 9789461738110 - 180
    solanum tuberosum - aardappelen - genomica - metabolisme - genetische analyse - metabolomica - metabolomen - fenotypen - loci voor kwantitatief kenmerk - solanum tuberosum - potatoes - genomics - metabolism - genetic analysis - metabolomics - metabolomes - phenotypes - quantitative trait loci

    Primary metabolism is essential for plant growth and survival and it is therefore involved in all physiological processes of the plant. In the past years the advancements in large-scale and high-throughput technologies have enhanced our ability to characterize the plant metabolome. The development of methods for the simultaneous analysis of many different plant metabolites and the necessary software for subsequent data analysis have further expanded the possibilities to investigate plant responses from a system-oriented perspective. This allows the comparison of genetic and phenotypic variation at different molecular levels, enabling us to find associations between genotype and phenotype and their intermediate levels of information transduction. Metabolomics has become increasingly important for the characterization of the metabolic status of plants under different environmental and genetic perturbations. The economic importance of potato and the increasing availability of genetic and molecular resources have stimulated research on many different aspects of the physiology of this crop and the regulation of complex traits. We used the available tools to explore the genetic basis of the composition and content of primary metabolites in a potato population. In this research, the possibilities to combine metabolite profiling with genetic information are explored to identify the genetic factors determining primary metabolism and to infer links between metabolites and agronomic phenotypes.

    Optimizing genomic selection for scarcely recorded traits
    Pszczola, M.J. - \ 2013
    Wageningen University. Promotor(en): Johan van Arendonk, co-promotor(en): Mario Calus; T. Strabel. - Wageningen : Wageningen UR - ISBN 9789461737663 - 158
    melkvee - genomen - selectief fokken - genetische verbetering - fokwaarde - fenotypen - genotypen - kenmerken - voeropname - dierveredeling - dairy cattle - genomes - selective breeding - genetic improvement - breeding value - phenotypes - genotypes - traits - feed intake - animal breeding

    Animal breeding aims to genetically improve animal populations by selecting the best individuals as parents of the next generation. New traits are being introduced to breeding goals to satisfy new demands faced by livestock production. Selecting for novel traits is especially challenging when recording is laborious and expensive and large scale recording is not possible. Genetic improvement of novel traits may be thus limited due to the small number of observations. New breeding tools, such as genomic selection, are therefore needed to enable the genetic improvement of novel traits. Using the limited available data optimally may, however, require alternative approaches and methodologies than currently used for conventional breeding goal traits. The overall objective of this thesis was to investigate different options for optimizing genomic selection for scarcely recorded novel traits. The investigated options were: (1) genotype imputation for ungenotyped but phenotyped animals to be used to enlarge the reference population; (2) optimization of the design of the reference population with respect to the relationships among the animals included in it; (3) prioritizing genotyping of the reference population or the selection candidates; and (4) using easily recordable predictor traits to improve the accuracy of breeding values for scarcely recorded traits. Results showed that: (1) including ungenotyped animals to the reference population can lead to a limited increase in the breeding values accuracy; (2) the reference population is designed optimally when the relationship within the reference are minimized and between reference population and potential selection candidates maximized; (3) the main gain in accuracy when moving from traditional to genomic selection is due to genotyping the selection candidates, but preferably both reference population and selection candidates should be genotyped; and (4) including the predictor traits in the analysis when it is recorded on both reference population and selection candidates can lead to a significant increase in the selection accuracy. The key factors for successful implementation of selection for a novel trait in a breeding scheme are: (1) maximizing accuracy of genotype prediction for ungenotyped animals to be used for updating the reference population; (2) optimizing the design of the reference population; (3) determining easy to record indicator traits that are also available on the selection candidates (4) developing large scale phenotyping techniques; and (5) establishing strategies and policies for increasing the engagement of farmers in the recording of novel traits.

    The ecology of life history evolution : genes, individuals and populations
    Visser, M.E. - \ 2013
    Wageningen : Wageningen University, Wageningen UR - ISBN 9789461735973
    ecologie - evolutie - genetica - natuurlijke selectie - fenotypen - populatie-ecologie - levensgeschiedenis - ecology - evolution - genetics - natural selection - phenotypes - population ecology - life history
    Natural selection shapes the life histories of organisms. The ecological interactions of these organisms with their biotic and abiotic environment shape the selection pressure on their phenotypes while their genetics determine how fast this selection leads to adaptation to their environment. The field of ecological genetics studies the response to natural selection in the wild and thus plays a key role in our understanding of the adaptive capacity of life, essential to understand how a changing environment affects the natural world.
    Genes for seed quality : integrating physiology and genetical genomics to mine for seed quality genes in tomato
    Kazmi, R.H. - \ 2013
    Wageningen University. Promotor(en): Harro Bouwmeester, co-promotor(en): Henk Hilhorst; Wilco Ligterink. - S.l. : s.n. - ISBN 9789461735201 - 243
    solanum lycopersicum - solanum pimpinellifolium - tomaten - zaadkwaliteit - genen - plantenfysiologie - genomica - fenotypen - metabolomica - solanum lycopersicum - solanum pimpinellifolium - tomatoes - seed quality - genes - plant physiology - genomics - phenotypes - metabolomics

    Seed quality in tomato is associated with many complex physiological and genetical traits. The performance of seeds is determined by three interlinked and interactive components that constitute a performance triangle of genetics, physiological quality and the environment. So far, there has been little or no discussion about the genetic analysis of seed and seedling traits in tomato at a systems level. To the best of our knowledge, the present study is the first systemic analysis of the genetics of seed and seedling traits, adding to a growing body of information on tomato seed quality. With the aim of improving the production of high-quality tomato seeds, a multidisciplinary study (physiology, genetics and genomics) was undertaken to develop and evaluate methods for improving the percentage, rate and uniformity of germination and early seedling development, and for increasing the range of environmental conditions for germination. Primarily, we explored the natural variation present in a Solanum lycopersicum x Solanum pimpinellifolium RIL population to dissect the molecular-genetic mechanisms controlling seed quality. Although previous solutions to issues associated with seed quality phenotypes seemed promising, none have utilized the integration of genomic, phenotypic and metabolic datasets to understand seed quality in tomato.Thus, the integration of metabolic and genomic analysis contributed to a comprehensive biological understanding of observed phenotypic differences between RILs of S. lycopersicumx S. pimpinellifolium. Here we describe, for the first time, the use of a generalized genetical genomics (GGG) model in tomato seeds that incorporates genetics, as well as environmental effects, and we applied this approach to map traditional quantitative trait loci (Genetic QTLs) and QTLs that are the result of interaction between the genetics and environmental changes (Genetic x Environmental QTLs). This model uses chosen environmental perturbations (different seed developmental stages, i.e. dry and 6h imbibed seeds) in combination with the analysis of genetic variation present in the RIL population, to study the change of metabolites over the multiple environments and to identify genotype-by-environment interactions. This thesis gives an account of the integration of genotyping, phenotyping and a molecular phenotype using metabolomics in generating a novel understanding of seed phenotypes and their interaction with the environment. In summary, the integration of phenotypic and metabolomics data has facilitated the identification of potential biomarkers for better understanding of the complex nature of tomato seed quality.

    Discovery and genotyping of existing and induced DNA sequence variation in potato
    Uitdewilligen, J.G.A.M.L. - \ 2012
    Wageningen University. Promotor(en): Richard Visser, co-promotor(en): Herman van Eck; Anne-Marie Wolters. - S.l. : s.n. - ISBN 9789461732330 - 165
    solanum tuberosum - aardappelen - dna-sequencing - dna - nucleotidenvolgordes - genotyping - plantenveredeling - genotypen - fenotypen - tetraploïdie - solanum tuberosum - potatoes - dna sequencing - dna - nucleotide sequences - genotyping - plant breeding - genotypes - phenotypes - tetraploidy

    In this thesis natural and induced DNA sequence diversity in potato (Solanum tuberosum) for use in marker-trait analysis and potato breeding is assessed. The study addresses the challenges of reliable, high-throughput identification and genotyping of sequence variants in existing tetraploid potato cultivar panels using traditional Sanger sequencing and next-generation massively parallel sequencing (MPS), and the application of this knowledge in the form of genetic markers. Furthermore, it explores the efficiency of ethyl methanesulphonate (EMS) mutagenesis combined with high resolution melting (HRM) DNA screening to induce and discover novel sequence variants in potato genotypes.
    Discovery and genotyping of sequence diversity in outcrossing autotetraploid species like potato is complex. In autotetraploid species, genotyping implies the quantitative identification of five alternative allele copy number states. In Chapter 1, several methodologies to identify and genotype DNA sequence variants, and the application of these sequence variants is discussed. This chapter provides an introduction to genotyping-by-sequencing (GBS) and the determination of allele copy number.
    In Chapter 2 the sequence diversity in three genes of the carotenoid pathway is assessed in diploid and tetraploid potato genotypes using direct Sanger sequencing. To investigate the genetics and molecular biology of orange and yellow flesh colour in potato, association analysis between SNP haplotypes and flesh colour phenotypes was performed, and the inheritance and gene expression of associated alleles was studied. We observed among eleven beta-carotene hydroxylase 2 (CHY2) alleles one dominant allele with a major effect, changing white into yellow flesh colour. In contrast, none of the lycopene epsilon cyclase (LCYe) alleles seemed to have a large effect on flesh colour. Analysis of zeaxanthin epoxidase (ZEP) alleles showed that a recessive allele with a non-LTR retrotransposon sequence in intron 1 reduced the expression level of the ZEP gene and caused accumulation of zeaxanthin. Genotypes combining presence of the dominant CHY2 allele with homozygosity for the recessive ZEP allele produced orange-fleshed tubers that accumulate large amounts of zeaxanthin.
    Sanger amplicon sequencing was applied in Chapter 3 to evaluate the sequence diversity in α-Glucan Water Dikinase (StGWD), a candidate gene underlying a QTL involved in potato starch phosphate content. Sanger sequences of two StGWD amplicons from a global collection of 398 commercial cultivars and progenitor lines were used to identify 16 unique haplotypes. By assigning tag SNPs to these haplotypes and by determining the allele copy number of identified sequence variants, we inferred the four-allele genetic composition for almost all cultivars assayed at this locus. This allowed genetic diversity parameters like the average number of different alleles present in a single cultivar (Ai=3.1) and the average intra-individual heterozygosity (Ho=0.765) to be estimated for this locus. Pedigree analysis confirmed that the identified haplotypes are identical by descent (IBD) and offered insight in the breeding history of elite potato germplasm. Haplotype association analysis led to the identification of two StGWD alleles causing altered starch phosphate content, which was further verified in diploid and tetraploid mapping populations containing the relevant alleles. One of these alleles (Allel H) increases the fraction of starch that is phosphorylated, while the other one (Allele A) decreases it.
    To scale up the discovery and genotyping of sequence variants, and to make it more whole-genome oriented, Chapter 4 reports on massively parallel sequencing (MPS) of approximately 800 genes scattered over the potato genome and resequenced in 83 tetraploid potato cultivars and a monoploid reference accession. We show that by combining MPS with genome complexity reduction and indexed sequencing, sufficient read depth for GBS can be achieved for reliable discovery and genotyping of sequence variants in individual tetraploid potato genotypes. With a custom designed, SureSelect enrichment library, 1.44 Mb of DNA sequence was targeted. The genes targeted were mainly single-copy genes, selected based on putative gene functions in both primary and secondary metabolic pathways, potato quality traits and biotic and abiotic stresses, and included a large set of conserved orthologous sequence genes (COSII) useful for genetic anchoring and phylogenetic studies. The indexed and enriched DNA libraries were sequenced on a Illumina HiSeq. After filtering and processing the raw sequence data, 12.4 Gb of high-quality sequence data was mapped to the potato genome, covering 2.1 Mb of the genome sequence with a median average read depth of 63× per cultivar. We detected over 129,000 sequence variants in these data and determined allele copy number of the variants in individual potato samples. The accuracy of the sequence-based allele copy number estimates was verified by a low-density SNP genotyping assay. This showed that for reliable genotyping a read count-based genotype quality score is best applied and a read depth of 80× is recommended for determining allele copy number in autotetraploid potato. Average nucleotide diversity (π=10.7×10-3 genome-wide, ≈1 variant/93 bp between two random alleles) varied along the twelve potato chromosomes, and individual genes under selection were identified. As an example for application of GBS for genome-wide association analysis (GWAS), the identified sequence variants and genotype data were tested in a marker-trait association analysis with plant maturity and tuber flesh colour. This led to the identification of alleles accounting for significant phenotypic variation in these traits.
    In Chapter 5 we applied the chemical mutagen EMS to diploid potato by two different treatments, a pollen and a seed treatment. We screened the resulting populations for novel mutations using HRM analysis. A pollen treatment with EMS dissolved in a sucrose solution was found to induce mutations only at a low frequency (only one mutation discovered after screening >2.7 Mb of sequence). In planta selection of the most vital mutagenized pollen seems to have lowered the mutation density to a frequency that is not suitable for reverse genetics studies. Treatment of potato seeds with EMS on the other hand provided a high density of novel mutations (1 mutation/65 kb), discovered in the M1 generation. In contrast to most EMS mutagenesis studies, we directly screened the M1 generation of the seed-treated population. In six candidate genes involved in potato starch and frying quality traits, 65 novel sequence variants were discovered. In all six genes, missense mutations that are predicted to damage protein function were discovered, and for four genes five premature stop codon mutations were identified. We attempted to stabilize and transfer 27 putatively interesting mutations to the M2 and M3 generation for further evaluation. Genetically stable M2 and M3 plants have been generated for 10 (37%) of these mutations. The estimated density of M1 mutations that are transferable to the M2 generation (one “accessible” mutation/118-176 kb) is higher than the mutation densities obtained in most other plant species, for which the M2 generation has been screened. The results of this chapter thus demonstrate that screening the M1 generation offers a good alternative to the commonly applied M2 screening for the rapid generation of novel genetic variation at a high density, without too much complication in recovering mutations in the M2 generation.
    In the concluding Chapter 6, results of preceding chapters are evaluated, and the prospects of the findings for potato research and breeding are discussed.

    Phenotypic changes in different spinach varieties grown and selected under organic conditions
    Serpolay, E. ; Schermann, N. ; Dawson, J.C. ; Lammerts Van Bueren, E. ; Goldringer, I. ; Chable, V. - \ 2011
    Sustainability 3 (2011)9. - ISSN 2071-1050 - p. 1616 - 1636.
    rassen (planten) - plantenvermeerdering - fenotypische selectie - fenotypen - spinacia oleracea - spinazie - conservering op het bedrijf - plantenveredeling - biologische landbouw - varieties - propagation - phenotypic selection - phenotypes - spinacia oleracea - spinach - on-farm conservation - plant breeding - organic farming
    Organic and low-input agriculture needs flexible varieties that can buffer environmental stress and adapt to the needs of farmers. We implemented an experiment to investigate the evolutionary capacities of a sample of spinach (Spinacia oleracea L.) population varieties for a number of phenotypic traits. Three farmers cultivated, selected and multiplied one or several populations over two years on their farms. The third year, the versions of the varieties cultivated and selected by the different farmers were compared to the original seed lots they had been given. After two cycles of cultivation and on-farm mass selection, all the observed varieties showed significant phenotypic changes (differences between the original version and the version cultivated by farmers) for morphological and phenological traits. When the divergence among versions within varieties was studied, the results show that the varieties conserved their identity, except for one variety, which evolved in such a way that it may now be considered two different varieties. The heterogeneity of the population varieties was assessed in comparison with a commercial F1 hybrid used as control, and we found no specific differences in phenotypic diversity between the hybrid and population varieties. The phenotypic changes shown by the population varieties in response to on-farm cultivation and selection could be useful for the development of specific adaptation. These results call into question the current European seed legislation and the requirements of phenotypic stability for conservation varieties
    The genetic architecture of gene expression in Caenorhabditis elegans
    Viñuela Rodriguez, A. - \ 2011
    Wageningen University. Promotor(en): Jaap Bakker, co-promotor(en): Jan Kammenga. - [S.l.] : S.n. - ISBN 9789085858386 - 119
    caenorhabditis elegans - genexpressie - stressfactoren - genetische analyse - transcriptie - loci voor kwantitatief kenmerk - genotypen - fenotypen - genexpressieanalyse - polygene overerving - caenorhabditis elegans - gene expression - stress factors - genetic analysis - transcription - quantitative trait loci - genotypes - phenotypes - genomics - polygenic inheritance

    Most organisms are exposed to a continuously changing environment throughout their life. For instance the ambient temperature is usually not constant and many species are exposed to a diverse range of anthropogenic stressors like toxic compounds. Moreover, individuals are prone to genetic changes due to mutation and allelic recombinations. All these factors might affect particular phenotypes, while others remain unchanged. This thesis provides insight into how phenotypic traits are affected by external stress factors and allelic recombinations in the nematode Caenorhabditis elegans (Nematoda; Rhabditidae). Because phenotypes and their variation may be explained by variation in gene expression, this thesis explored the architecture of gene expression and some of the elements that contribute to gene expression.

    Chapters 2 and 3 focus on environmental stressors with i) a specific target (two organophosphorus pesticides) and ii) a non-target mode of action (temperature) to study their influence on gene expression. A single genotype, the canonical wild type strain Bristol (N2) was used to study the effect of interacting pesticides by exposing nematodes to a toxicant mixture at two different temperatures Analysis revealed common transcriptional responses related to detoxification, stress, innate immunity, and transport of lipids to all treatments. It was found that for both pesticides these similar processes were regulated by different gene transcripts in single and combined treatments. These results also showed that the effect of a mix of low doses of pesticides is not a summed effect of the single components. Moreover, increased temperature elevates the toxic consequences to the pesticides exposures. This toxicity gain is attributed to an elevated uptake and accumulation of the toxicants in the organisms. These results support the idea that the observed higher toxicity of pesticides with temperature might be a consequence of gene-environment interactions affecting detoxification genes. Together, thefirst part of this thesis illustrates the intense crosstalk between gene pathways in response to interacting environmental stressors in C. elegans.

    The second part of the thesis elaborates on the influence of different genotypes as multiple perturbations on gene expression. How the genotype-phenotype relationship progresses with age was investigated using a quantitative genetics approach (genetical genomics). We performed a genetic mapping strategy of gene transcription variation (expression-QTL, eQTL) to explore the dynamics of regulatory loci affecting genome-wide gene expression at three different ages. We used a recombinant inbred line (RIL) population generated from a cross between the C. elegans strain N2 and the wild type CB4856 in Chapter 4. Also, we investigated the influence of age to reveal a genotype-by-age effect (gxaeQTL) on gene expression. The total number of detected eQTL decreased with age whereas the variation in expression increased. In developing worms, the number of genes with increased expression variation (1282) was similar to the ones with decreased expression variation (1328). In aging worms the number of genes with increased variation (1772) was nearly 5 times higher than the number of genes with a decreased expression variation (373). Furthermore, the number of cis-acting eQTL in juveniles decreased by almost 50% in old worms whereas the number of trans-acting loci decreased by ~27%, indicating that cis-regulation becomes relatively less frequent than trans-regulation in aging worms. Our findings demonstrate that eQTL patterns are strongly affected by age and suggest that gene network integrity declines with age. To better understand the changes in the gene network with age, gene expression profiles of N2 and CB4856 were generated for Chapter 5. We explored gene expression heritability and transgression as genetic parameters for the analysis of gene expression divergence in different genotypes. The average broad sense heritability was similar in developing and aging worms; but the gene expression variance that can be attributed to genetic variance in each gene changes with age. It can be proposed that regulation became more polygenic in aging worms. These changes explain the decrease in detected eQTLs. Likewise, it explains the imbalance between highly heritable genes and eQTLs in aging worms.

    Chapter 6 discusses the main conclusion of this thesis in the context of the robustness theory. Robustness in biological systems provides the potential to survive severe environmental and genetic perturbations in the form of cryptic genetic variation. The variation we observed in gene transcripts due to external and internal perturbations not always translated to physiological phenotypic variation. In some cases however, the mechanisms underlying phenotypic robustness failed and phenotypic variation was observed. Such genetic cryptic variation was revealed as new molecular and physiological phenotypes.

    Agrobacterium-mediated transformation of Mycosphaerella fijiensis, the devastating Black Sigatoka pathogen of bananas
    Díaz-Trujillo, C. ; Adibon, H. ; Kobayashi, K. ; Zwiers, L.H. ; Souza, M.T. ; Kema, G.H.J. - \ 2010
    Gewasbescherming 41 (2010)3. - ISSN 0166-6495 - p. 151 - 151.
    mycosphaerella fijiensis - fungiciden - bananen - genotypen - fenotypen - rhizobium - genetische transformatie - genoomanalyse - mycosphaerella fijiensis - fungicides - bananas - genotypes - phenotypes - rhizobium - genetic transformation - genome analysis
    Mycosphaerella fijiensis, M. musicola en M. eumusae veroorzaken de Sigatoka-ziekte in banaan. Op dit moment is de toepassing van fungiciden de enige optie om deze ziekte te bestrijden. Het PRPB (Pesticide Reduction Program for Banana) investeert in de ontwikkeling van technieken voor de genotype- en fenotypebepaling van M. fijiensis. Hierbij wordt gebruikt gemaakt van ATMT (Agrobacterium tumefaciens-mediated transformation).
    Microsatellite genotyping of apple (Malus × domestica Borkh.) genetic resources in the Netherlands: application in collection management and variety identification
    Treuren, R. van; Kemp, H. ; Ernsting, G. ; Jongejans, B. ; Houtman, H. ; Visser, L. - \ 2010
    Genetic Resources and Crop Evolution 57 (2010)6. - ISSN 0925-9864 - p. 853 - 865.
    appels - genenbanken - genotypen - genetische merkers - microsatellieten - genetische diversiteit - fenotypen - apples - gene banks - genotypes - genetic markers - microsatellites - genetic diversity - phenotypes - molecular characterization - ssr markers - construction - polymorphism - database - tomato - plants - dna
    A highly informative set of 16 microsatellite markers was used to fingerprint 695 apple accessions from eight Dutch collections. Among the total sample, 475 different genotypes were distinguished based on multi-locus microsatellite variation, revealing a potential redundancy within the total sample of 32%. The majority of redundancies were found between collections, rather than within collections. No single collection covered the total observed diversity well, as each collection consisted of about 50% of unique accessions. These findings reflected the fact that most collection holders focus on common Dutch varieties, as well as on region-specific diversity. Based on the diversity patterns observed, maintenance of genetic resources by a network of co-operating collection holders, rather than by collecting the total diversity in a single collection appears to be an efficient approach. Comparison of microsatellite and passport data showed that for many accessions the marker data did not provide support for the registered variety names. Verification of accessions showed that discrepancies between passport and molecular data were largely due to documentation and phenotypic determination errors. With the help of the marker data the varietal names of 45 accessions could be corrected. Microsatellite genotyping of apple appears to be an efficient tool in the management of collections and in variety identification. The development of a marker database was considered relevant as a reference instrument in variety identification and as a source of information about thus far unexplored diversity that could be of interest in the development of new apple varieties
    Phytophthora: kampioen aanpassing
    Kessel, G.J.T. ; Evenhuis, A. ; Bosch, G.B.M. van den; Förch, M.G. ; Topper, C.G. - \ 2009
    phytophthora infestans - genotypen - fenotypen - virulentie - fungiciden - aardappelen - ziekteresistentie - phytophthora infestans - genotypes - phenotypes - virulence - fungicides - potatoes - disease resistance
    Poster met onderzoeksinformatie. Uit onbeschermd opgekweekte proefveldjes zijn Phytophthora infestans isolaten verzameld en gekarakteriseerd.
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