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"Thomson, J. Michael"
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An improved 7K SNP array, the C7AIR, provides a wealth of validated SNP markers for rice breeding and genetics studies
Single nucleotide polymorphisms (SNPs) are highly abundant, amendable to high-throughput genotyping, and useful for a number of breeding and genetics applications in crops. SNP frequencies vary depending on the species and populations under study, and therefore target SNPs need to be carefully selected to be informative for each application. While multiple SNP genotyping systems are available for rice (Oryza sativa L. and its relatives), they vary in their informativeness, cost, marker density, speed, flexibility, and data quality. In this study, we report the development and performance of the Cornell-IR LD Rice Array (C7AIR), a second-generation SNP array containing 7,098 markers that improves upon the previously released C6AIR. The C7AIR is designed to detect genome-wide polymorphisms within and between subpopulations of O. sativa, as well as O. glaberrima, O. rufipogon and O. nivara. The C7AIR combines top-performing SNPs from several previous rice arrays, including 4,007 SNPs from the C6AIR, 2,056 SNPs from the High Density Rice Array (HDRA), 910 SNPs from the 384-SNP GoldenGate sets, 189 SNPs from the 44K array selected to add information content for elite U.S. tropical japonica rice varieties, and 8 trait-specific SNPs. To demonstrate its utility, we carried out a genome-wide association analysis for plant height, employing the C7AIR across a diversity panel of 189 rice accessions and identified 20 QTLs contributing to plant height. The C7AIR SNP chip has so far been used for genotyping >10,000 rice samples. It successfully differentiates the five subpopulations of Oryza sativa, identifies introgressions from wild and exotic relatives, and is useful for quantitative trait loci (QTL) and association mapping in diverse materials. Moreover, data from the C7AIR provides valuable information that can be used to select informative and reliable SNP markers for conversion to lower-cost genotyping platforms for genomic selection and other downstream applications in breeding.
Journal Article
Optimization of gene editing in cowpea through protoplast transformation and agroinfiltration by targeting the phytoene desaturase gene
Cowpea ( Vigna unguiculata ) is a legume staple widely grown across Sub-Saharan Africa and other tropical and sub-tropical regions. Considering projected climate change and global population increases, cowpea’s adaptation to hot climates, resistance to drought, and nitrogen-fixing capabilities make it an especially attractive crop for facing future challenges. Despite these beneficial traits, efficient varietal improvement is challenging in cowpea due to its recalcitrance to transformation and long regeneration times. Transient gene expression assays can provide solutions to alleviate these issues as they allow researchers to test gene editing constructs before investing in the time and resource- intensive process of transformation. In this study, we developed an improved cowpea protoplast isolation protocol, a transient protoplast assay, and an agroinfiltration assay to be used for initial testing and validation of gene editing constructs and for gene expression studies. To test these protocols, we assessed the efficacy of a CRISPR-Cas9 construct containing four multiplexed single-guide RNA (sgRNA) sequences using polyethylene glycol (PEG)-mediated transformation and agroinfiltration with phytoene desaturase ( PDS ) as the target gene. Sanger sequencing of DNA from transformed protoplasts and agroinfiltrated cowpea leaves revealed several large deletions in the target sequences. The protoplast system and agroinfiltration protocol developed in this study provide versatile tools to test gene editing components before initiating plant transformation, thus improving the chance of using active sgRNAs and attaining the desired edits and target phenotype.
Journal Article
EcoEnsemble: A general framework for combining ecosystem models in R
Often there are several complex ecosystem models available to address a specific question. However, structural differences, systematic discrepancies and uncertainties mean that they typically produce different outputs. Rather than selecting a single ‘best’ model, it is desirable to combine them to give a coherent answer to the question at hand. Many methods of combining ecosystem models assume that one of the models is exactly correct, which is unlikely to be the case. Furthermore, models may not be fitted to the same data, have the same outputs, nor be run for the same time period, making many common methods difficult to implement. In this paper, we use a statistical model to describe the relationship between the ecosystem models, prior beliefs and observations to make coherent predictions of the true state of the ecosystem with robust quantification of uncertainty. We introduce EcoEnsemble, an R package that takes advantage of the statistical model's structure to efficiently fit the ensemble model, either sampling from the posterior distribution or maximising the posterior density. We demonstrate EcoEnsemble by investigating what would happen to four fish species in the North Sea under future management scenarios. Although developed for applications in ecology, EcoEnsemble can be used to combine any group of mechanistic models, for example in climate modelling, epidemiology or biology.
Journal Article
Genetic Control of Plasticity in Root Morphology and Anatomy of Rice in Response to Water Deficit
Elucidating the genetic control of rooting behavior under water-deficit stress is essential to breed climate-robust rice (Oryza sativa) cultivars. Using a diverse panel of 274 indica genotypes grown under control and water-deficit conditions during vegetative growth, we phenotyped 35 traits, mostly related to root morphology and anatomy, involving 45,000 root-scanning images and nearly 25,000 cross sections from the root-shoot junction. The phenotypic plasticity of these traits was quantified as the relative change in trait value under water-deficit compared with control conditions. We then carried out a genome-wide association analysis on these traits and their plasticity, using 45,608 high-quality single-nucleotide polymorphisms. One hundred four significant loci were detected for these traits under control conditions, 106 were detected under water-deficit stress, and 76 were detected for trait plasticity. We predicted 296 (control), 284 (water-deficit stress), and 233 (plasticity) a priori candidate genes within linkage disequilibrium blocks for these loci. We identified key a priori candidate genes regulating root growth and development and relevant alleles that, upon validation, can help improve rice adaptation to water-deficit stress.
Journal Article
Exploring novel genetic sources of salinity tolerance in rice through molecular and physiological characterization
Agricultural productivity is increasingly being affected by the build-up of salinity in soils and water worldwide. The genetic base of salt-tolerant rice donors being used in breeding is relatively narrow and needs broadening to breed varieties with wider adaptation to salt-affected areas. This study evaluated a large set of rice accessions of diverse origins to identify and characterize novel sources of salt tolerance.
Diversity analysis was performed on 107 germplasm accessions using a genome-wide set of 376 single-nucleotide polymorphism (SNP) markers, along with characterization of allelic diversity at the major quantitative trait locus Saltol Sixty-nine accessions were further evaluated for physiological traits likely associated with responses to salt stress during the seedling stage.
Three major clusters corresponding to the indica, aus and aromatic subgroups were identified. The largest group was indica, with the salt-tolerant Pokkali accessions in one sub-cluster, while a set of Bangladeshi landraces, including Akundi, Ashfal, Capsule, Chikirampatnai and Kutipatnai, were in a different sub-cluster. A distinct aus group close to indica contained the salt-tolerant landrace Kalarata, while a separate aromatic group closer to japonica rice contained a number of traditional, but salt-sensitive Bangladeshi landraces. These accessions have different alleles at the Saltol locus. Seven landraces - Akundi, Ashfal, Capsule, Chikirampatnai, Jatai Balam, Kalarata and Kutipatnai - accumulated less Na and relatively more K, maintaining a lower Na/K ratio in leaves. They effectively limit sodium transport to the shoot.
New salt-tolerant landraces were identified that are genetically and physiologically distinct from known donors. These landraces can be used to develop better salt-tolerant varieties and could provide new sources of quantitative trait loci/alleles for salt tolerance for use in molecular breeding. The diversity observed within this set and in other donors suggests multiple mechanisms that can be combined for higher salt tolerance.
Journal Article
Transcriptome profiling of two rice varieties reveals their molecular responses under high night-time temperature
High night-time temperatures (HNT) pose a threat to the sustainability of crop production, including rice. HNT can affect crop productivity and quality by influencing plant physiology, morphology, and phenology. The ethylene perception inhibitor, 1-methylcyclopropene (1-MCP), can minimize HNT-induced damage to plant membranes, thereby preventing decrease in rice yield. In this study, we employed a transcriptome approach to investigate the effects of HNT, 1-MCP, and their interaction on two Texas rice varieties, Antonio and Colorado. The plants were exposed to temperatures of 25°C (ambient night-time temperature, ANT) and 30°C (HNT) using an infrared heating system from the booting stage until harvest, while 1-MCP was applied at the booting stage of rice development. Several physiological and agronomical traits were evaluated under each condition to assess plant responses. Leaf tissues were collected from the plants grown in the ANT and HNT conditions after the heat stress and 1-MCP treatments. Based on agronomic performance, Colorado was less negatively affected than Antonio under HNT, showing a slight reduction in spikelet fertility and leaf photosynthetic rate but no significant reduction in yield. The application of 1-MCP significantly mitigated the adverse effects of HNT in Antonio. However, no significant differences were observed in yield and leaf photosynthetic rate in Colorado. Furthermore, transcriptomic data revealed distinct responsive mechanisms in Antonio and Colorado in response to both HNT and 1-MCP. Several ethylene and senescence-related transcription factors (TFs) were identified only in Antonio, suggesting that 1-MCP affected the ethylene signaling pathway in Antonio but not in Colorado. These findings contribute to our understanding of the physiological differences between varieties exhibiting susceptible and tolerant responses to high night-time temperatures, as well as their response to 1-MCP and ethylene regulation under 1-MCP.
Journal Article
Genetic and genomic approaches to develop rice germplasm for problem soils
Soils that contain toxic amounts of minerals or are deficient in essential plant nutrients are widespread globally and seriously constrain rice production. New methods are necessary to incorporate the complex adaptive traits associated with tolerance of these abiotic stresses, while simultaneously retaining the high yield potential of rice varieties when conditions are favorable. Significant progress in the genetic characterization of stress response pathways and recent advances in genomics have provided powerful tools for in-depth dissection of tolerance mechanisms. Additionally, tolerance of most of these abiotic stresses in rice is controlled by a few QTLs with large effects despite the intricacy of the numerous traits involved. Genetic dissection of these QTLs and their incorporation into high-yielding varieties will significantly enhance and stabilize rice productivity in these problem soils. Current efforts at IRRI and in rice breeding programs worldwide are seeking to explore diverse germplasm collections and genetically dissect the causal mechanisms of tolerance to facilitate their use in breeding. This review focuses on salinity and P and Zn deficiency as the major problems encountered in rice soils, and examines current understanding of the mechanisms involved and efforts toward germplasm improvement.
Journal Article
The role of microRNA-1 and microRNA-133 in skeletal muscle proliferation and differentiation
Understanding the molecular mechanisms that regulate cellular proliferation and differentiation is a central theme of developmental biology. MicroRNAs (miRNAs) are a class of regulatory RNAs of ∼22 nucleotides that post-transcriptionally regulate gene expression
1
,
2
. Increasing evidence points to the potential role of miRNAs in various biological processes
3
,
4
,
5
,
6
,
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,
8
. Here we show that miRNA-1 (miR-1) and miRNA-133 (miR-133), which are clustered on the same chromosomal loci, are transcribed together in a tissue-specific manner during development. miR-1 and miR-133 have distinct roles in modulating skeletal muscle proliferation and differentiation in cultured myoblasts
in vitro
and in
Xenopus laevis
embryos
in vivo
. miR-1 promotes myogenesis by targeting histone deacetylase 4 (HDAC4), a transcriptional repressor of muscle gene expression. By contrast, miR-133 enhances myoblast proliferation by repressing serum response factor (SRF). Our results show that two mature miRNAs, derived from the same miRNA polycistron and transcribed together, can carry out distinct biological functions. Together, our studies suggest a molecular mechanism in which miRNAs participate in transcriptional circuits that control skeletal muscle gene expression and embryonic development.
Journal Article
MicroRNAs Regulate Brain Morphogenesis in Zebrafish
MicroRNAs (miRNAs) are small RNAs that regulate gene expression posttranscriptionally. To block all miRNA formation in zebrafish, we generated maternal-zygotic dicer (MZdicer) mutants that disrupt the Dicer ribonuclease III and double-stranded RNA-binding domains. Mutant embryos do not process precursor miRNAs into mature miRNAs, but injection of preprocessed miRNAs restores gene silencing, indicating that the disrupted domains are dispensable for later steps in silencing. MZdicer mutants undergo axis formation and differentiate multiple cell types but display abnormal morphogenesis during gastrulation, brain formation, somitogenesis, and heart development. Injection of miR-430 miRNAs rescues the brain defects in MZdicer mutants, revealing essential roles for miRNAs during morphogenesis.
Journal Article