MarkerMiner 1.0: a new application for phylogenetic marker development using angiosperm transcriptomes

Premise of the study: Targeted sequencing using next-generation sequencing (NGS) platforms offers enormous potential for plant systematics by enabling economical acquisition of multilocus data sets that can resolve difficult phylogenetic problems. However, because discovery of single-copy nuclear (SCN) loci from NGS data requires both bioinformatics skills and access to high-performance computing resources, the application of NGS data has been limited. Methods and Results: We developed MarkerMiner 1.0, a fully automated, open-access bioinformatic workflow and application for discovery of SCN loci in angiosperms. Our new tool identified as many as 1993 SCN loci from transcriptomic data sampled as part of four independent test cases representing marker development projects at different phylogenetic scales. Conclusions: MarkerMiner is an easy-to-use and effective tool for discovery of putative SCN loci. It can be run locally or via the Web, and its tabular and alignment outputs facilitate efficient downstream assessments of phylogenetic utility, locus selection, intron-exon boundary prediction, and primer or probe development

Validation of reference transcripts in strawberry (<i>Fragaria</i> spp.)

Contemporary methods to assay gene expression depend on a stable set of reference transcripts for accurate quantitation. A lack of well-tested reference genes slows progress in characterizing gene expression in high-value specialty crops. In this study, a set of strawberry (Fragaria spp.) constitutively expressed reference genes has been identified by merging digital gene expression data with expression profiling. Constitutive reference candidates were validated using quantitative PCR and hybridization. Several transcripts have been identified that show improved stability across tissues relative to traditional reference transcripts. Results are similar between commercial octoploid strawberry and the diploid model. Our findings also show that while some never-before-used references are appropriate for most applications, even the most stable reference transcripts require careful assessment across the diverse tissues and fruit developmental states before being adopted as controls.Facultad de Ciencias ExactasInstituto de Fisiología Vegeta

The mammalian LINC complex regulates genome transcriptional responses to substrate rigidity

Mechanical integration of the nucleus with the extracellular matrix (ECM) is established by linkage between the cytoskeleton and the nucleus. This integration is hypothesized to mediate sensing of ECM rigidity, but parsing the function of nucleus-cytoskeleton linkage from other mechanisms has remained a central challenge. Here we took advantage of the fact that the LINC (linker of nucleoskeleton and cytoskeleton) complex is a known molecular linker of the nucleus to the cytoskeleton, and asked how it regulates the sensitivity of genome-wide transcription to substratum rigidity. We show that gene mechanosensitivity is preserved after LINC disruption, but reversed in direction. Combined with myosin inhibition studies, we identify genes that depend on nuclear tension for their regulation. We also show that LINC disruption does not attenuate nuclear shape sensitivity to substrate rigidity. Our results show for the first time that the LINC complex facilitates mechano-regulation of expression across the genome

Integrative analyses of TEDDY Omics data reveal lipid metabolism abnormalities, increased intracellular ROS and heightened inflammation prior to autoimmunity for type 1 diabetes

27 páginas, 6 figuras, Contiene material suplementarioBackground: The Environmental Determinants of Diabetes in the Young (TEDDY) is a prospective birth cohort designed to study type 1 diabetes (T1D) by following children with high genetic risk. An integrative multi-omics approach was used to evaluate islet autoimmunity etiology, identify disease biomarkers, and understand progression over time. Results: We identify a multi-omics signature that was predictive of islet autoimmunity (IA) as early as 1 year before seroconversion. At this time, abnormalities in lipid metabolism, decreased capacity for nutrient absorption, and intracellular ROS accumulation are detected in children progressing towards IA. Additionally, extracellular matrix remodeling, inflammation, cytotoxicity, angiogenesis, and increased activity of antigen-presenting cells are observed, which may contribute to beta cell destruction. Our results indicate that altered molecular homeostasis is present in IA-developing children months before the actual detection of islet autoantibodies, which opens an interesting window of opportunity for therapeutic intervention. Conclusions: The approach employed herein for assessment of the TEDDY cohort showcases the utilization of multi-omics data for the modeling of complex, multifactorial diseases, like T1D.This study was supported by grant 2015PG-T1D050 provided by the Leona M. and Harry B. Helmsley Charitable Trust. The TEDDY Study is funded by U01 DK63829, U01 DK63861, U01 DK63821, U01 DK63865, U01 DK63863, U01 DK63836, U01 DK63790, UC4 DK63829, UC4 DK63861, UC4 DK63821, UC4 DK63865, UC4 DK63863, UC4 DK63836, UC4 DK95300, UC4 DK100238, UC4 DK106955, UC4 DK112243, UC4 DK117483, and Contract No. HHSN267200700014C from the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), National Institute of Allergy and Infectious Diseases (NIAID), National Institute of Child Health and Human Development (NICHD), National Institute of Environmental Health Sciences (NIEHS), Centers for Disease Control and Prevention (CDC), and JDRF. This work was supported in part by the NIH/NCATS Clinical and Translational Science Awards to the University of Florida (UL1 TR000064) and the University of Colorado (UL1 TR001082).Peer reviewe

A Consensus Genetic Map for Pinus taeda and Pinus elliottii and Extent of Linkage Disequilibrium in Two Genotype-Phenotype Discovery Populations of Pinus taeda

A consensus genetic map for Pinus taeda (loblolly pine) and Pinus elliottii (slash pine) was constructed by merging three previously published P. taeda maps with a map from a pseudo-backcross between P. elliottii and P. taeda. The consensus map positioned 3856 markers via genotyping of 1251 individuals from four pedigrees. It is the densest linkage map for a conifer to date. Average marker spacing was 0.6 cM and total map length was 2305 cM. Functional predictions of mapped genes were improved by aligning expressed sequence tags used for marker discovery to full-length P. taeda transcripts. Alignments to the P. taeda genome mapped 3305 scaffold sequences onto 12 linkage groups. The consensus genetic map was used to compare the genome-wide linkage disequilibrium in a population of distantly related P. taeda individuals (ADEPT2) used for association genetic studies and a multiple-family pedigree used for genomic selection (CCLONES). The prevalence and extent of LD was greater in CCLONES as compared to ADEPT2; however, extended LD with LGs or between LGs was rare in both populations. The average squared correlations, r2, between SNP alleles less than 1 cM apart were less than 0.05 in both populations and r2 did not decay substantially with genetic distance. The consensus map and analysis of linkage disequilibrium establish a foundation for comparative association mapping and genomic selection in P. taeda and P. elliottii.J.W.W. was supported by a USDA CSREES Food and Agricultural Sciences National Needs Graduate Fellowship. V.E.C. was supported by USDA NIFA Award #2011-68002-30185 (PINEMAP) and the USDA Forest Service. L.S.W. was supported by the National Science Foundation under grant no. ABI-1062432 to Indiana University. P.M.G., D.N., and K.M. were supported in part by USDA NIFA Award #2011-67009-30030 (PineRefSeq) to University of California, Davis.Peer reviewe

A physical map for the Amborella trichopoda genome sheds light on the evolution of angiosperm genome structure

Background: Recent phylogenetic analyses have identified Amborella trichopoda, an understory tree species endemic to the forests of New Caledonia, as sister to a clade including all other known flowering plant species. The Amborella genome is a unique reference for understanding the evolution of angiosperm genomes because it can serve as an outgroup to root comparative analyses. A physical map, BAC end sequences and sample shotgun sequences provide a first view of the 870 Mbp Amborella genome.Results: Analysis of Amborella BAC ends sequenced from each contig suggests that the density of long terminal repeat retrotransposons is negatively correlated with that of protein coding genes. Syntenic, presumably ancestral, gene blocks were identified in comparisons of the Amborella BAC contigs and the sequenced Arabidopsis thaliana, Populus trichocarpa, Vitis vinifera and Oryza sativa genomes. Parsimony mapping of the loss of synteny corroborates previous analyses suggesting that the rate of structural change has been more rapid on lineages leading to Arabidopsis and Oryza compared with lineages leading to Populus and Vitis. The gamma paleohexiploidy event identified in the Arabidopsis, Populus and Vitis genomes is shown to have occurred after the divergence of all other known angiosperms from the lineage leading to Amborella.Conclusions: When placed in the context of a physical map, BAC end sequences representing just 5.4% of the Amborella genome have facilitated reconstruction of gene blocks that existed in the last common ancestor of all flowering plants. The Amborella genome is an invaluable reference for inferences concerning the ancestral angiosperm and subsequent genome evolution

Maintaining informatics training learning outcomes with a COVID-19 era shift to a fully online flipped course

The emergence of the coronavirus disease 2019 pandemic forced us to adapt our recently developed informatics training serving a variety of students as well as faculty and staff. The successful flipped classroom course series (a hybrid-format with both asynchronous online learning and in-person synchronous components) was shifted to a fully online format with the synchronous portion now held via web-based video conference. We repeated our participant survey at the end of each of the 3 one-credit courses to compare student satisfaction and learning outcomes achievement to the original offering. The responses were overall very positive again with a slight response distribution improvement in some measures of satisfaction. Likewise, students reported similar achievement of the learning outcomes across all courses with some Unix coourse objectives receiving higher competency agreement in the new, fully online version. Overall, the fully online version of the course series was equally successful, if not more so, than the original version with a physical classroom session each week. Given that participants also had strong agreement with a new question that they would prefer online class meetings instead of in a classroom, even if there wasn’t a global pandemic (citing a variety of logistical reasons such as “convenience of screen sharing,” parking issues, and job-related time constraints), the fully online version of the informatics training will be retained

Primers for Low-Copy Nuclear Genes in <i>Metrosideros</i> and Cross-Amplification in Myrtaceae

Premise of the study: Primers were developed to amplify low-copy nuclear genes in Hawaiian Metrosideros (Myrtaceae). Methods and Results: Data from a pooled 454 Titanium run of the partial transcriptomes of four Metrosideros taxa were used to identify the loci of interest. Ten exon-primed intron-crossing (EPIC) markers were amplified and sequenced directly with success in Metrosideros, as well as in a representative selection of Myrtaceae, including Syzygium, Psidium, and Melaleuca for most of the markers. The loci amplified ranged between 500 and 1100 bp, and up to 117 polymorphic sites were observed within an individual gene alignment. Two introns contained microsatellites in some of the species. Conclusions: These novel primer pairs should be useful for phylogenetic analysis and population genetics of a broad range of Myrtaceae, particularly the diverse fleshy-fruited tribes Syzygieae and Myrteae

Primers for Low-Copy Nuclear Genes in the Hawaiian Endemic <i>Clermontia</i> (Campanulaceae) and Cross-Amplification in Lobelioideae

Premise of the study: Primers were developed to amplify 12 intron-less, low-copy nuclear genes in the Hawaiian genus <i>Clermontia</i> (Campanulaceae), a suspected tetraploid. Methods and Results: Data from a pooled 454 titanium run of the partial transcriptomes of seven <i>Clermontia</i> species were used to identify the loci of interest. Most loci were amplified and sequenced directly with success in a representative selection of lobeliads even though several of these loci turned out to be duplicated. Levels of variation were comparable to those observed in commonly used plastid and ribosomal markers. Conclusions: We found evidence of a genome duplication that likely predates the diversification of the Hawaiian lobeliads. Some genes nevertheless appear to be single-copy and should be useful for phylogenetic studies of <i>Clermontia</i> or the entire Lobelioideae subfamily