RESUMO
Meiosis is key to sexual reproduction and genetic diversity. Here, we show that the Arabidopsis cyclin-dependent kinase Cdk1/Cdk2 homolog CDKA;1 is an important regulator of meiosis needed for several aspects of meiosis such as chromosome synapsis. We identify the chromosome axis protein ASYNAPTIC 1 (ASY1), the Arabidopsis homolog of Hop1 (homolog pairing 1), essential for synaptonemal complex formation, as a target of CDKA;1. The phosphorylation of ASY1 is required for its recruitment to the chromosome axis via ASYNAPTIC 3 (ASY3), the Arabidopsis reductional division 1 (Red1) homolog, counteracting the disassembly activity of the AAA+ ATPase PACHYTENE CHECKPOINT 2 (PCH2). Furthermore, we have identified the closure motif in ASY1, typical for HORMA domain proteins, and provide evidence that the phosphorylation of ASY1 regulates the putative self-polymerization of ASY1 along the chromosome axis. Hence, the phosphorylation of ASY1 by CDKA;1 appears to be a two-pronged mechanism to initiate chromosome axis formation in meiosis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/citologia , Arabidopsis/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Proteínas de Ligação a DNA/metabolismo , Meiose , Adenosina Trifosfatases/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Sítios de Ligação , Cromossomos de Plantas/genética , Cromossomos de Plantas/metabolismo , Quinases Ciclina-Dependentes/genética , Proteínas de Ligação a DNA/química , Mutação , Fosforilação , Ligação Proteica , Multimerização ProteicaRESUMO
KEY MESSAGE: Multiple QTLs control unreduced pollen production in potato. Two major-effect QTLs co-locate with mutant alleles of genes with homology to AtJAS, a known regulator of meiotic spindle orientation. In diploid potato the production of unreduced gametes with a diploid (2n) rather than a haploid (n) number of chromosomes has been widely reported. Besides their evolutionary important role in sexual polyploidisation, unreduced gametes also have a practical value for potato breeding as a bridge between diploid and tetraploid germplasm. Although early articles argued for a monogenic recessive inheritance, the genetic basis of unreduced pollen production in potato has remained elusive. Here, three diploid full-sib populations were genotyped with an amplicon sequencing approach and phenotyped for unreduced pollen production across two growing seasons. We identified two minor-effect and three major-effect QTLs regulating this trait. The two QTLs with the largest effect displayed a recessive inheritance and an additive interaction. Both QTLs co-localised with genes encoding for putative AtJAS homologs, a key regulator of meiosis II spindle orientation in Arabidopsis thaliana. The function of these candidate genes is consistent with the cytological phenotype of mis-oriented metaphase II plates observed in the parental clones. The alleles associated with elevated levels of unreduced pollen showed deleterious mutation events: an exonic transposon insert causing a premature stop, and an amino acid change within a highly conserved domain. Taken together, our findings shed light on the natural variation underlying unreduced pollen production in potato and will facilitate interploidy breeding by enabling marker-assisted selection for this trait.
Assuntos
Arabidopsis , Solanum tuberosum , Melhoramento Vegetal , Pólen/genética , Genótipo , Arabidopsis/genética , MeioseRESUMO
Chromosome substitution lines (CSLs) are tentatively supreme resources to investigate non-allelic genetic interactions. However, the difficulty of generating such lines in most species largely yielded imperfect CSL panels, prohibiting a systematic dissection of epistasis. Here, we present the development and use of a unique and complete panel of CSLs in Arabidopsis thaliana, allowing the full factorial analysis of epistatic interactions. A first comparison of reciprocal single chromosome substitutions revealed a dependency of QTL detection on different genetic backgrounds. The subsequent analysis of the complete panel of CSLs enabled the mapping of the genetic interactors and identified multiple two- and three-way interactions for different traits. Some of the detected epistatic effects were as large as any observed main effect, illustrating the impact of epistasis on quantitative trait variation. We, therefore, have demonstrated the high power of detection and mapping of genome-wide epistasis, confirming the assumed supremacy of comprehensive CSL sets.
Assuntos
Arabidopsis , Mapeamento Cromossômico , Cromossomos de Plantas , Epistasia Genética , Locos de Características Quantitativas , Arabidopsis/genética , Mapeamento Cromossômico/métodos , Cromossomos de Plantas/genética , Genoma de Planta , FenótipoRESUMO
Through the inactivation of genes that act during meiosis it is possible to direct the genetic make-up of plants in subsequent generations and optimize breeding schemes. Offspring may show higher recombination of parental alleles resulting from elevated crossover (CO) incidence, or by omission of meiotic divisions, offspring may become polyploid. However, stable mutations in genes essential for recombination, or for either one of the two meiotic divisions, can have pleiotropic effects on plant morphology and line stability, for instance by causing lower fertility. Therefore, it is often favorable to temporarily change gene expression during meiosis rather than relying on stable null mutants. It was previously shown that virus-induced gene silencing (VIGS) can be used to transiently reduce CO frequencies. We asked if VIGS could also be used to modify other processes throughout meiosis and during pollen formation in Arabidopsis thaliana. Here, we show that VIGS-mediated knock-down of FIGL1, RECQ4A/B, OSD1 and QRT2 can induce (i) an increase in chiasma numbers, (ii) unreduced gametes and (iii) pollen tetrads. We further show that VIGS can target both sexes and different genetic backgrounds and can simultaneously silence different gene copies. The successful knock-down of these genes in A. thaliana suggests that VIGS can be exploited to manipulate any process during or shortly after meiosis. Hence, the transient induction of changes in inheritance patterns can be used as a powerful tool for applied research and biotechnological applications.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , ATPases Associadas a Diversas Atividades Celulares/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Expressão Gênica , Inativação Gênica , Meiose/genética , Proteínas Associadas aos Microtúbulos/genética , Melhoramento Vegetal , Pólen/genética , Pólen/metabolismoRESUMO
Little is known how patterns of cross-over (CO) numbers and distribution during meiosis are established. Here, we reveal that cyclin-dependent kinase A;1 (CDKA;1), the homolog of human Cdk1 and Cdk2, is a major regulator of meiotic recombination in ArabidopsisArabidopsis plants with reduced CDKA;1 activity experienced a decrease of class I COs, especially lowering recombination rates in centromere-proximal regions. Interestingly, this reduction of type I CO did not affect CO assurance, a mechanism by which each chromosome receives at least one CO, resulting in all chromosomes exhibiting similar genetic lengths in weak loss-of-function cdka;1 mutants. Conversely, an increase of CDKA;1 activity resulted in elevated recombination frequencies. Thus, modulation of CDKA;1 kinase activity affects the number and placement of COs along the chromosome axis in a dose-dependent manner.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Quinases Ciclina-Dependentes/fisiologia , Recombinação Genética , Alelos , Arabidopsis/citologia , Proteínas de Arabidopsis/fisiologia , Cromossomos de Plantas , Troca Genética , MeioseRESUMO
Plant breeding applications exploiting meiotic mutant phenotypes (like the increase or decrease of crossover (CO) recombination) have been proposed over the last years. As recessive meiotic mutations in breeding lines may affect fertility or have other pleiotropic effects, transient silencing techniques may be preferred. Reverse breeding is a breeding technique that would benefit from the transient downregulation of CO formation. The technique is essentially the opposite of plant hybridization: a method to extract parental lines from a hybrid. The method can also be used to efficiently generate chromosome substitution lines (CSLs). For successful reverse breeding, the two homologous chromosome sets of a heterozygous plant must be divided over two haploid complements, which can be achieved by the suppression of meiotic CO recombination and the subsequent production of doubled haploid plants. Here we show the feasibility of transiently reducing CO formation using virus-induced gene silencing (VIGS) by targeting the meiotic gene MSH5 in a wild-type heterozygote of Arabidopsis thaliana. The application of VIGS (rather than using lengthy stable transformation) generates transgene-free offspring with the desired genetic composition: we obtained parental lines from a wild-type heterozygous F1 in two generations. In addition, we obtained 20 (of the 32 possible) CSLs in one experiment. Our results demonstrate that meiosis can be modulated at will in A. thaliana to generate CSLs and parental lines rapidly for hybrid breeding. Furthermore, we illustrate how the modification of meiosis using VIGS can open routes to develop efficient plant breeding strategies.
Assuntos
Arabidopsis/genética , Segregação de Cromossomos , Inativação Gênica , Melhoramento Vegetal/métodos , Proteínas de Arabidopsis/genética , Quimera , Cromossomos de Plantas , Troca Genética , Proteínas de Ligação a DNA/genética , Fertilidade/genética , Regulação da Expressão Gênica de Plantas , Meiose , Mutação , Fenótipo , TransgenesRESUMO
Meiosis is a specialized eukaryotic cell division that generates haploid gametes required for sexual reproduction. During meiosis, homologous chromosomes pair and undergo reciprocal genetic exchange, termed crossover (CO). Meiotic CO frequency varies along the physical length of chromosomes and is determined by hierarchical mechanisms, including epigenetic organization, for example methylation of the DNA and histones. Here we investigate the role of DNA methylation in determining patterns of CO frequency along Arabidopsis thaliana chromosomes. In A. thaliana the pericentromeric regions are repetitive, densely DNA methylated, and suppressed for both RNA polymerase-II transcription and CO frequency. DNA hypomethylated methyltransferase1 (met1) mutants show transcriptional reactivation of repetitive sequences in the pericentromeres, which we demonstrate is coupled to extensive remodeling of CO frequency. We observe elevated centromere-proximal COs in met1, coincident with pericentromeric decreases and distal increases. Importantly, total numbers of CO events are similar between wild type and met1, suggesting a role for interference and homeostasis in CO remodeling. To understand recombination distributions at a finer scale we generated CO frequency maps close to the telomere of chromosome 3 in wild type and demonstrate an elevated recombination topology in met1. Using a pollen-typing strategy we have identified an intergenic nucleosome-free CO hotspot 3a, and we demonstrate that it undergoes increased recombination activity in met1. We hypothesize that modulation of 3a activity is caused by CO remodeling driven by elevated centromeric COs. These data demonstrate how regional epigenetic organization can pattern recombination frequency along eukaryotic chromosomes.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , DNA (Citosina-5-)-Metiltransferases/genética , DNA de Plantas/genética , Epigenômica , Meiose/genética , Recombinação Genética , Proteínas de Arabidopsis/metabolismo , Centrômero , Cromossomos de Plantas/química , Cromossomos de Plantas/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA , DNA Intergênico , DNA de Plantas/metabolismo , Histonas/genética , Histonas/metabolismo , Mutação , Mapeamento Físico do Cromossomo , Sequências Repetitivas de Ácido Nucleico , TelômeroRESUMO
BACKGROUND: Solanum dulcamara (bittersweet, climbing nightshade) is one of the few species of the Solanaceae family native to Europe. As a common weed it is adapted to a wide range of ecological niches and it has long been recognized as one of the alternative hosts for pathogens and pests responsible for many important diseases in potato, such as Phytophthora. At the same time, it may represent an alternative source of resistance genes against these diseases. Despite its unique ecology and potential as a genetic resource, genomic research tools are lacking for S. dulcamara. We have taken advantage of next-generation sequencing to speed up research on and use of this non-model species. RESULTS: In this work, we present the first large-scale characterization of the S. dulcamara transcriptome. Through comparison of RNAseq reads from two different accessions, we were able to predict transcript-based SNP and SSR markers. Using the SNP markers in combination with genomic AFLP and CAPS markers, the first genome-wide genetic linkage map of bittersweet was generated. Based on gene orthology, the markers were anchored to the genome of related Solanum species (tomato, potato and eggplant), revealing both conserved and novel chromosomal rearrangements. This allowed a better estimation of the evolutionary moment of rearrangements in a number of cases and showed that chromosomal breakpoints are regularly re-used. CONCLUSION: Knowledge and tools developed as part of this study pave the way for future genomic research and exploitation of this wild Solanum species. The transcriptome assembly represents a resource for functional analysis of genes underlying interesting biological and agronomical traits and, in the absence of the full genome, provides a reference for RNAseq gene expression profiling aimed at understanding the unique biology of S. dulcamara. Cross-species orthology-based marker selection is shown to be a powerful tool to quickly generate a comparative genetic map, which may speed up gene mapping and contribute to the understanding of genome evolution within the Solanaceae family.
Assuntos
Genômica , Solanum/genética , Cromossomos de Plantas/genética , Análise por Conglomerados , Evolução Molecular , Perfilação da Expressão Gênica , Marcadores Genéticos/genética , Repetições de Microssatélites/genética , Modelos Genéticos , Anotação de Sequência Molecular , Polimorfismo de Nucleotídeo Único/genética , Especificidade da EspécieRESUMO
Plants can express different phenotypic responses following polyploidization, but ploidy-dependent phenotypic variation has so far not been assigned to specific genetic factors. To map such effects, segregating populations at different ploidy levels are required. The availability of an efficient haploid inducer line in Arabidopsis thaliana allows for the rapid development of large populations of segregating haploid offspring. Because Arabidopsis haploids can be self-fertilised to give rise to homozygous doubled haploids, the same genotypes can be phenotyped at both the haploid and diploid ploidy level. Here, we compared the phenotypes of recombinant haploid and diploid offspring derived from a cross between two late flowering accessions to map genotype × ploidy (G × P) interactions. Ploidy-specific quantitative trait loci (QTLs) were detected at both ploidy levels. This implies that mapping power will increase when phenotypic measurements of monoploids are included in QTL analyses. A multi-trait analysis further revealed pleiotropic effects for a number of the ploidy-specific QTLs as well as opposite effects at different ploidy levels for general QTLs. Taken together, we provide evidence of genetic variation between different Arabidopsis accessions being causal for dissimilarities in phenotypic responses to altered ploidy levels, revealing a G × P effect. Additionally, by investigating a population derived from late flowering accessions, we revealed a major vernalisation-specific QTL for variation in flowering time, countering the historical bias of research in early flowering accessions.
Assuntos
Arabidopsis , Mapeamento Cromossômico , Genótipo , Locos de Características Quantitativas/genética , HaploidiaRESUMO
Chromosomal rearrangements are relatively rare evolutionary events and can be used as markers to study karyotype evolution. This research aims to use such rearrangements to study chromosome evolution in Solanum. Chromosomal rearrangements between Solanum crops and several related wild species were investigated using tomato and potato bacterial artificial chromosomes (BACs) in a multicolour fluorescent in situ hybridization (FISH). The BACs selected are evenly distributed over seven chromosomal arms containing inversions described in previous studies. The presence/absence of these inversions among the studied Solanum species were determined and the order of the BAC-FISH signals was used to construct phylogenetic trees.Compared with earlier studies, data from this study provide support for the current grouping of species into different sections within Solanum; however, there are a few notable exceptions, such as the tree positions of S. etuberosum (closer to the tomato group than to the potato group) and S. lycopersicoides (sister to S. pennellii). These apparent contradictions might be explained by interspecific hybridization events and/or incomplete lineage sorting. This cross-species BAC painting technique provides unique information on genome organization, evolution and phylogenetic relationships in a wide variety of species. Such information is very helpful for introgressive breeding.
Assuntos
Cromossomos de Plantas/genética , Evolução Molecular , Genoma de Planta , Hibridização in Situ Fluorescente/métodos , Solanum/genética , Inversão Cromossômica/genética , Cromossomos Artificiais Bacterianos/genética , Cruzamentos Genéticos , Endogamia , Cariótipo , Cariotipagem/métodos , Filogenia , Solanum/classificação , Especificidade da Espécie , SinteniaRESUMO
Crossover recombination is a crucial process in plant breeding because it allows plant breeders to create novel allele combnations on chromosomes that can be used for breeding superior F1 hybrids. Gaining control over this process, in terms of increasing crossover incidence, altering crossover positions on chromosomes or silencing crossover formation, is essential for plant breeders to effectively engineer the allelic composition of chromosomes. We review the various means of crossover control that have been described or proposed. By doing so, we sketch a field of science that uses both knowledge from classic literature and the newest discoveries to manage the occurrence of crossovers for a variety of breeding purposes.
Assuntos
Cruzamento/métodos , Cromossomos de Plantas , Troca Genética , Plantas/genética , Genótipo , MeioseRESUMO
Clonal propagation and genetic engineering of plants requires regeneration, but many species are recalcitrant and there is large variability in explant responses. Here, we perform a genome-wide association study using 190 natural Arabidopsis accessions to dissect the genetics of shoot regeneration from root explants and several related in vitro traits. Strong variation is found in the recorded phenotypes and association mapping pinpoints a myriad of quantitative trait genes, including prior candidates and potential novel regeneration determinants. As most of these genes are trait- and protocol-specific, we propose a model wherein shoot regeneration is governed by many conditional fine-tuning factors and a few universal master regulators such as WUSCHEL, whose transcript levels correlate with natural variation in regenerated shoot numbers. Potentially novel genes in this last category are AT3G09925, SUP, EDA40 and DOF4.4. We urge future research in the field to consider multiple conditions and genetic backgrounds.
Assuntos
Arabidopsis/genética , Brotos de Planta/fisiologia , Regeneração/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/genética , Variação Genética/genética , Estudo de Associação Genômica Ampla , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/fisiologia , Reguladores de Crescimento de Plantas/genética , Locos de Características Quantitativas/genética , Característica Quantitativa HerdávelRESUMO
Assessment of the impact of variation in chloroplast and mitochondrial DNA (collectively termed the plasmotype) on plant phenotypes is challenging due to the difficulty in separating their effect from nuclear-derived variation (the nucleotype). Haploid-inducer lines can be used as efficient plasmotype donors to generate new plasmotype-nucleotype combinations (cybrids)1. We generated a panel comprising all possible cybrids of seven Arabidopsis thaliana accessions and extensively phenotyped these lines for 1,859 phenotypes under both stable and fluctuating conditions. We show that natural variation in the plasmotype results in both additive and epistatic effects across all phenotypic categories. Plasmotypes that induce more additive phenotypic changes also cause more epistatic effects, suggesting a possible common basis for both additive and epistatic effects. On average, epistatic interactions explained twice as much of the variance in phenotypes as additive plasmotype effects. The impact of plasmotypic variation was also more pronounced under fluctuating and stressful environmental conditions. Thus, the phenotypic impact of variation in plasmotypes is the outcome of multi-level nucleotype-plasmotype-environment interactions and, as such, the plasmotype is likely to serve as a reservoir of variation that is predominantly exposed under certain conditions. The production of cybrids using haploid inducers is a rapid and precise method for assessment of the phenotypic effects of natural variation in organellar genomes. It will facilitate efficient screening of unique nucleotype-plasmotype combinations to both improve our understanding of natural variation in these combinations and identify favourable combinations to enhance plant performance.
Assuntos
Arabidopsis/genética , Genoma de Planta , Organelas/genética , Fenótipo , Hibridização GenéticaRESUMO
Reverse breeding (RB) is a novel plant breeding technique designed to directly produce parental lines for any heterozygous plant, one of the most sought after goals in plant breeding. RB generates perfectly complementing homozygous parental lines through engineered meiosis. The method is based on reducing genetic recombination in the selected heterozygote by eliminating meiotic crossing over. Male or female spores obtained from such plants contain combinations of non-recombinant parental chromosomes which can be cultured in vitro to generate homozygous doubled haploid plants (DHs). From these DHs, complementary parents can be selected and used to reconstitute the heterozygote in perpetuity. Since the fixation of unknown heterozygous genotypes is impossible in traditional plant breeding, RB could fundamentally change future plant breeding. In this review, we discuss various other applications of RB, including breeding per chromosome.
Assuntos
Cruzamento/métodos , Técnicas de Silenciamento de Genes , Meiose , Desenvolvimento Vegetal , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Troca Genética , Genoma de Planta , Heterozigoto , Plantas/genéticaRESUMO
To follow the dynamics of meiosis in the model plant Arabidopsis, we have established a live cell imaging setup to observe male meiocytes. Our method is based on the concomitant visualization of microtubules (MTs) and a meiotic cohesin subunit that allows following five cellular parameters: cell shape, MT array, nucleus position, nucleolus position, and chromatin condensation. We find that the states of these parameters are not randomly associated and identify 11 cellular states, referred to as landmarks, which occur much more frequently than closely related ones, indicating that they are convergence points during meiotic progression. As a first application of our system, we revisited a previously identified mutant in the meiotic A-type cyclin TARDY ASYNCHRONOUS MEIOSIS (TAM). Our imaging system enabled us to reveal both qualitatively and quantitatively altered landmarks in tam, foremost the formation of previously not recognized ectopic spindle- or phragmoplast-like structures that arise without attachment to chromosomes.
Assuntos
Arabidopsis/citologia , Arabidopsis/crescimento & desenvolvimento , Microscopia Intravital/métodos , Meiose , Células Vegetais/fisiologia , Organelas/metabolismo , Organelas/ultraestrutura , Células Vegetais/químicaRESUMO
High-quality chromosome images of mitotic and meiotic cell divisions in plant tissues are inextricably connected with the technical control of cell spread preparations. Superb chromosome slides are the best for studying chromosome morphology and making karyotypes; they also are the best start for a successful fluorescent in situ hybridization experiment. In this study, we describe the essentials for fixation, enzymatic digestion, squash, spread, and dropping protocols for clean and well-differentiated nuclei and chromosome complements. In addition, we focus on the use of standard whole image processing for best sharpness, brightness and contrast adjustments, differentiation of heterochromatin/euchromatin, and high dynamic range imaging of big chromosomes. We also explain how to combine transparent layers or spot channels of different fluorescent images for making publication quality, full color photos.
Assuntos
Cromossomos de Plantas/genética , Heterocromatina/genética , Eucromatina/genética , Hibridização In Situ , CariotipagemRESUMO
To produce seeds, flowering plants need to specify somatic cells to undergo meiosis. Here, we reveal a regulatory cascade that controls the entry into meiosis starting with a group of redundantly acting cyclin-dependent kinase (CDK) inhibitors of the KIP-RELATED PROTEIN (KRP) class. KRPs function by restricting CDKA;1-dependent inactivation of the Arabidopsis Retinoblastoma homolog RBR1. In rbr1 and krp triple mutants, designated meiocytes undergo several mitotic divisions, resulting in the formation of supernumerary meiocytes that give rise to multiple reproductive units per future seed. One function of RBR1 is the direct repression of the stem cell factor WUSCHEL (WUS), which ectopically accumulates in meiocytes of triple krp and rbr1 mutants. Depleting WUS in rbr1 mutants restored the formation of only a single meiocyte.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/embriologia , Proteínas de Homeodomínio/metabolismo , Óvulo Vegetal/embriologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas de Ciclo Celular , Proteínas Inibidoras de Quinase Dependente de Ciclina/genética , Proteínas Inibidoras de Quinase Dependente de Ciclina/metabolismo , Proteínas de Homeodomínio/genética , Meiose/genética , Meiose/fisiologia , Mutação , Óvulo Vegetal/genética , Óvulo Vegetal/metabolismoRESUMO
Meiosis is a key event of sexual life cycles in eukaryotes. Its mechanistic details have been uncovered in several model organisms, and most of its essential features have received various and often contradictory evolutionary interpretations. In this perspective, we present an overview of these often 'weird' features. We discuss the origin of meiosis (origin of ploidy reduction and recombination, two-step meiosis), its secondary modifications (in polyploids or asexuals, inverted meiosis), its importance in punctuating life cycles (meiotic arrests, epigenetic resetting, meiotic asymmetry, meiotic fairness) and features associated with recombination (disjunction constraints, heterochiasmy, crossover interference and hotspots). We present the various evolutionary scenarios and selective pressures that have been proposed to account for these features, and we highlight that their evolutionary significance often remains largely mysterious. Resolving these mysteries will likely provide decisive steps towards understanding why sex and recombination are found in the majority of eukaryotes.This article is part of the themed issue 'Weird sex: the underappreciated diversity of sexual reproduction'.
Assuntos
Evolução Biológica , Eucariotos/fisiologia , Meiose , Recombinação Genética , SexoRESUMO
INTRODUCTION: Batch effects in large untargeted metabolomics experiments are almost unavoidable, especially when sensitive detection techniques like mass spectrometry (MS) are employed. In order to obtain peak intensities that are comparable across all batches, corrections need to be performed. Since non-detects, i.e., signals with an intensity too low to be detected with certainty, are common in metabolomics studies, the batch correction methods need to take these into account. OBJECTIVES: This paper aims to compare several batch correction methods, and investigates the effect of different strategies for handling non-detects. METHODS: Batch correction methods usually consist of regression models, possibly also accounting for trends within batches. To fit these models quality control samples (QCs), injected at regular intervals, can be used. Also study samples can be used, provided that the injection order is properly randomized. Normalization methods, not using information on batch labels or injection order, can correct for batch effects as well. Introducing two easy-to-use quality criteria, we assess the merits of these batch correction strategies using three large LC-MS and GC-MS data sets of samples from Arabidopsis thaliana. RESULTS: The three data sets have very different characteristics, leading to clearly distinct behaviour of the batch correction strategies studied. Explicit inclusion of information on batch and injection order in general leads to very good corrections; when enough QCs are available, also general normalization approaches perform well. Several approaches are shown to be able to handle non-detects-replacing them with very small numbers such as zero seems the worst of the approaches considered. CONCLUSION: The use of quality control samples for batch correction leads to good results when enough QCs are available. If an experiment is properly set up, batch correction using the study samples usually leads to a similar high-quality correction, but has the advantage that more metabolites are corrected. The strategy for handling non-detects is important: choosing small values like zero can lead to suboptimal batch corrections.
RESUMO
Hybrid crop varieties are traditionally produced by selecting and crossing parental lines to evaluate hybrid performance. Reverse breeding allows doing the opposite: selecting uncharacterized heterozygotes and generating parental lines from them. With these, the selected heterozygotes can be recreated as F1 hybrids, greatly increasing the number of hybrids that can be screened in breeding programs. Key to reverse breeding is the suppression of meiotic crossovers in a hybrid plant to ensure the transmission of nonrecombinant chromosomes to haploid gametes. These gametes are subsequently regenerated as doubled-haploid (DH) offspring. Each DH carries combinations of its parental chromosomes, and complementing pairs can be crossed to reconstitute the initial hybrid. Achiasmatic meiosis and haploid generation result in uncommon phenotypes among offspring owing to chromosome number variation. We describe how these features can be dealt with during a reverse-breeding experiment, which can be completed in six generations (â¼1 year).