Browsing by Subject "genome"

Now showing 1 - 5 of 5
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    Comparative genomics approaches accurately predict deleterious variants in plants
    (2018-07-03) Kono, Thomas John Y; Lei, Li; Shih, Ching-Hua; Hoffman, Paul J; Morrell, Peter L; Fay, Justin C; pmorrell@umn.edu; Morrell, Peter L; University of Minnesota Department of Agronomy and Plant Genetics; University of Rochester Department of Biology
    Recent advances in genome resequencing have led to increased interest in prediction of the functional consequences of genetic variants. Variants at phylogenetically conserved sites are of particular interest, because they are more likely than variants at phylogenetically variable sites to have deleterious effects on fitness and contribute to phenotypic variation. Numerous comparative genomic approaches have been developed to predict deleterious variants, but they are nearly always judged based on their ability to identify known disease-causing mutations in humans. Determining the accuracy of deleterious variant predictions in nonhuman species is important to understanding evolution, domestication, and potentially to improving crop quality and yield. To examine our ability to predict deleterious variants in plants we generated a curated database of 2,910 Arabidopsis thaliana mutants with known phenotypes. We evaluated seven approaches and found that while all performed well, the single best-performing approach was a likelihood ratio test applied to homologs identified in 42 plant genomes. Although the approaches did not always agree, we found only slight differences in performance when comparing mutations with gross versus biochemical phenotypes, duplicated versus single copy genes, and when using a single approach versus ensemble predictions. We conclude that deleterious mutations can be reliably predicted in A. thaliana and likely other plant species, but that the relative performance of various approaches can depend on the organism to which they are applied.
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    DNA-based detection and reference genome assembly of Aphanomyces cochlioides
    (2021-08) Botkin , Jacob
    Aphanomyces root rot (ARR) and Aphanomyces damping-off, caused by the soil-borne oomycete A. cochlioides, are common diseases of sugar beet in major production regions. Management techniques are implemented to mitigate losses, but ultimately A. cochlioides is intractable and significantly reduces the sucrose content of the sugar beet taproot throughout the growing season, especially during periods with above average precipitation. Currently, a genome sequence for A. cochlioides is not available, and existing diagnostic assays are time consuming and have limitations. The first objective was to assemble and annotate a reference genome for A. cochlioides. We conducted a de novo genome assembly and annotation of A. cochlioides using 232x coverage of Nanopore long-reads, and error corrected with 77x coverage of Illumina short-reads. The assembled genome was 76.3 Mb, consisted of 97 contigs, and had a contig N50 of 2.6 Mb. The assembly contained 93.2% of complete benchmarking universal single-copy orthologs, a repeat content of 32.1%, and 20,274 gene models. This is the first report of a reference genome for A. cochlioides, which could serve as a platform for future investigations into virulence mechanisms, comparative genomics, and the development of diagnostic assays. The second objective was to develop a rapid, sensitive, and accurate DNA-based detection assay to quantify A. cochlioides inoculum in infested soil and infected sugar beet tissue. We developed a TaqMan qPCR assay that was specific to A. cochlioides. The qPCR assay was validated with 12 naturally infested soil samples, which had Ct (cycle threshold) values of 26.72 to 34.64 and ARR disease severity index (DSI) values of 48 to 100. The qPCR assay was further validated on infected adult sugar beets and seedlings. For 60 adult sugar beet roots, A. cochlioides DNA was detected in 63% of the samples, while a culture-based assay identified A. cochlioides in 15% of the samples. Furthermore, A. cochlioides DNA was detected in infected seedlings as early as 5 days after planting in a naturally infested soil. Finally, when oospore infested potting soil was tested with the qPCR assay and ARR bioassay, a strong correlation was observed between oospore density and Ct value (R2 = 0.96), as well as oospore density and DSI value (R2 = 0.968). The limit of detection (LOD) was 5 oospores per g soil (dry wt.), which had a mean Ct value of 34.58, and a mean DSI value of 23.33. Our DNA-based detection assay could provide growers with the A. cochlioides infestation level of field soils to help them make informed management decisions prior to planting.
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    Genomic resources to study virulence and evolution of cereal rust fungi
    (2021-05) Henningsen, Eva
    Stem rust caused by Puccinia graminis f. sp. tritici (Pgt) and crown rust caused by Pucciniacoronata f. sp. avenae (Pca) are global threats the production of wheat and oat, respectively. Fast evolving populations of both Pgt and Pca limit the efficacy of plant genetic resistance and constrain disease management strategies. Chapter 1 provides background information about both rust fungi and their biology, shares a comprehensive review of the available genome resources in the rusts, and highlights some advancements in rust research and how they can be utilized. Chapter 2 describes a study where my colleagues and I developed a pipeline for identifying candidate susceptibility genes for future study of stem rust virulence using comparative transcriptome-based and orthology-guided approaches. The analysis was targeted to genes with differential expression in T. aestivum and genes suppressed or not affected in B. distachyon and reports several processes potentially linked to susceptibility to Pgt, such as cell death suppression and impairment of photosynthesis. The approach was complemented with a gene co-expression network analysis to identify wheat targets to deliver resistance to Pgt through removal or modification of putative susceptibility genes. This work could help further the understanding of the molecular mechanisms that lead to rust infection and disease susceptibility; this in turn could deliver novel strategies to deploy crop resistance through genetic loss of disease susceptibility. A significant contribution of this work is a pipeline that can be adapted to study virulence of other rust fungi. Finally, Chapter 3 describes a high-quality genome assembly of Pca isolate 203. The ultimate goal of the assembly is to provide the first fully haplotype-phased, chromosome level reference for Pca. To this end, PacBio long reads and Illumina short reads were obtained to create the initial draft assembly, while Hi-C reads were collected to order contigs and phase the genome. Contigs were assigned to haplotype bins using gene synteny initially, and these bins were aligned to the Pgt 21-0 A haplotype genome to evaluate the probable number of chromosomes and possible chromosome sizes. Future steps for completing the high-quality assembly include an iterative process to fix haplotype phase swaps through manual curation and scaffolding, final chromosome assignment, and annotation with RNAseq data. A collection of publications with my contributions is provided in the appendix section.
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    Sequence analysis of a rice BAC covering the syntenous barley Rpg1 region
    (Genome, 1999) Steffenson, Brian; Han, F.; Kilian, A.; Chen, J.P.; Kudrna, D.; Yamamoto, K.; Matsumoto, T.; Sasaki, T.; Kleinhofs, A.
    In the course of map-based cloning of the barley stem rust resistance gene Rpg1, we identified a rice bacterial artificial chromosome (BAC) containing the Rpg1 flanking markers. Based on the excellent gene order colinearity between barley and rice in this region, we expected that this rice BAC would contain the barley Rpg1 homologue. In order to identify the putative rice homologue, we sequenced ca. 35 kb of the rice BAC at random and then an additional 33 kb of contiguous sequence between the two most closely spaced Rpg1 flanking markers. Sequence analysis revealed a total of 15 putative genes, 5 within the 33-kb contiguous region. A rice Rpg1 homologue was not identified, although a gene encoding a hypothetical polypeptide with similarity to a membrane protein could not be eliminated as a candidate. Surprisingly, four of the genes identified in the 33-kb contiguous rice sequence showed a high degree of similarity with genes on Arabidopsis chromosome 4. The genome regions harboring these genes showed some relatedness, but many rearrangements were also evident. These data suggest that some genes have remained linked even over the long evolutionary separation of Arabidopsis and rice, as has also been reported for mammals and invertebrates.
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    Whole Genome Imputation Panel of 624 Dogs
    (2023-03-09) Friedenberg, Steven; Clark, Leigh Anne; Murphy, Sarah; Greif, Elizabeth; Evans, Jacquelyn; Tsai, Kate; fried255@umn.edu; Friedenberg, Steven
    This dataset contains a compressed variant call file (VCF) and index file of phased, bi-allelic, single nucleotide variants (SNVs) from 624 dogs of various breeds that were used as a reference panel for imputation of low-pass whole-genome sequencing from 83 Great Danes. Also included is an Excel file containing breed information for each of the 624 dogs. The file contains data for all 38 canine autosomes and the X chromosome.