Project description:There is growing evidence for the prevalence of DNA copy number variation (CNV) and its role in phenotypic variation in recent years. Comparative genomic hybridization (CGH) was used to explore the extent of this type of structural variation in the barley genome. In a panel of 14 genotypes including domesticated cultivars and wild barleys, we found that 14.9% of all the sequences on the array are affected by CNV. Higher levels of CNV diversity are present in the wild accessions relative to cultivated barley. A substantial portion (37%) of the CNV events are present in both wild and domesticated barley. CNVs are enriched in telomeric regions for all chromosomes except 4H, which is also the barley chromosome with the lowest proportion of CNVs. CNV affected 9.5% of the coding sequences represented on the array. The genes affected by CNV are enriched for sequences annotated as disease-resistance proteins and protein kinases, suggesting the potential for CNV to influence variation for responses to biotic and abiotic stress. The analysis of CNV breakpoints indicated that DNA repair mechanisms of double-strand breaks (DSBs) via single-stranded annealing (SSA) and synthesis-dependent strand annealing (SDSA) play an important role in the origin of many structural changes in barley. Here we present the first catalog of CNVs in a diploid Triticeae species, which opens the door for future genome diversity research in a tribe that comprises the economically important cereal species wheat, barley and rye. Our findings constitute a valuable resource for the identification of CNV affecting genes of agronomic importance.
Project description:To evaluate the DNA methylation in LSK cells from the bone marrow of wildtype or Tet2/3 DKO mice. In order to address the impact of the loss of Tet2/3 proteins in DNA methylation in LSK cells, we compared by WGBS the methylome of wild and, Tet2/3 DKO LSK cells in bone marrow.
Project description:We hypothesized that the genome segments of cultivated barley should show certain similarity with its ancestral wild barley. Instead of whole genome sequences, we employed RNA-Seq to investigated the genomic origin of modern cultivated barley using some representative wild barley genotypes from the Near East and Tibet, and representative world-wide selections of cultivated barley.
Project description:Maintaining genome integrity presents a particular challenge for plants due to their sedentary lifestyle, which disables direct avoidance of unfavorable external conditions. Additionally, plants’ metabolic processes generate reactive molecules as by-products of e.g. photosynthesis, creating internal DNA-damaging conditions. Due to this, plants have developed a unique and strictly regulated web of DNA damage responses (DDR). We initiated analysis of the DDR system in cultivated barley (Hordeum vulgare), a temperate cereal model with a large and repeat rich genome. A series of DNA damaging assays was established to describe barley plants’ phenotypic response to chemically induced DNA double-strand breaks. The efficacy of assays as a tool to be used for assessing potential new DDR barley mutants was demonstrated. DNA damage response network activation in barley was assessed by transcriptome analysis using RNA sequencing for wild type and mutant in the DDR signaling kinase ATAXIA TELANGIECTASIA MUTATED AND RAD3-RELATED (ATR). Considering the barley genome had only recently been sequenced, and it lacks the in-depth gene analysis, a list of potential DNA damage response genes in barley was compiled based on their homology with Arabidopsis genes. The comparison of transcripts in wild-type plants and atr mutants following genotoxic stress
Project description:Drought is a destructive abiotic stress, with plants possessing diverse strategies to survive periods of limited water resources. Previous results have described connections between strigolactone (SL) and drought, however, here we used the barley (Hordeum vulgare) SL-insensitive mutant hvd14 (dwarf14) to investigate the SL-dependent mechanisms related to water deficit response. By combining transcriptome, proteome with phytohormone analyses and physiological data, we describe the drought-mediated differences between wild-type and hvd14 plants. Our findings indicate that the drought sensitivity of hvd14 is related to weaker induction of abscisic acid-responsive genes/proteins, lower jasmonic acid content, higher reactive oxygen species content, and lower wax biosynthic and deposition mechanisms then wild-type plants. In addition, we identify a series of transcription factors (TFs) that are exclusively drought-induced in wild-type barley. Critically, we resolve a comprehensive series interestions between the drought-induced barley transcriptome and proteome responses that allow us to understand the impacts of SL in mitigating water limiting conditions. These data provide a number of new angles for the development of drought-resistant barley.
Project description:The development of whole-genome bisulfite sequencing (WGBS) has led to a number of exciting discoveries about how genomes utilize DNA methylation and has led to a plethora of novel testable hypotheses. Methods for constructing sodium bisulfite-converted and amplified libraries have recently excelled to the point that the bottleneck for experiments that use WGBS has shifted to data analysis and interpretation. Here we present empirical evidence for an over-representation of methylated DNA from WGBS. This enrichment for methylated DNA is exacerbated by higher cycles of PCR and is influenced by the type of uracil-insensitive DNA polymerase used for amplifying the sequencing library. Future efforts to computationally correct for this enrichment bias will be essential to increasing the accuracy of determining methylation levels for individual cytosines. MethylC-Seq of Arabidopsis thaliana
Project description:We used unbiased whole genome bisulfite sequencing (WGBS) to identify DNA methylation changes in the intestinal stem cells (ISCs) or their progeny during the suckling period of mouse colon development. Lgr5-EGFP mice were used to identify ISC populations in the colons. WGBS were performed using EGFP labeled Lgr5+ ISCs and epithelial cell adhesion molecule (EpCAM) labeled epithelial cells isolated at the beginning and end of the suckling period (postnatal day 0-P0 and P21).