Project description:Our previous experiment on biogenesis mechanisim of stress-inducible antisense RNAs showed that accumulation of antisense RNA in a drought-inducible gene, RD29A locus was decreased in rdr1/2/6 triple mutant. The RD29A antisense RNA did not have CAP structure and poly(A) tail. We applied deep sequencing of uncapped RNA using SOLiD to identify RDR1/2/6-mediated antisense RNAs in genome-wide. We identified 1) 975 RDR1/2/6-mediated antisense RNA loci on which no. of sequence tags is decreased in rdr1/2/6 and makes peak and 2) 1,613 RDR1/2/6-mediated antisense RNA loci on which no. of the tags is decreased in rdr1/2/6 and enriched on antisense strand of exons. They were grouped into 1,959 non-redundant loci.
Project description:Identification of microRNA expressed in Arabidopsis plants grown at different ambient CO2 concentrations and different ambient temperatures
Project description:Transcriptional profiling of Arabidopsis thaliana 12-days old seedlings comparing Col-0 wild type with transgenic plants with altered expression of dual-targetting plastid/mitochondrial organellar RNA-polymerase RPOTmp. Transgenic plants used for experiment were: overexpressor plants obtained by transformation of Col-0 WT plants with genetic constructs created in [Tarasenko et al., 2016] contained catalytic part of RPOTmp enzyme with transit peptides of RPOTm (mitochondrial) and RPOTp (plastid) by agrobacterial transformation; plants with complementation of RPOTmp functions in mitochondria or chloroplasts obtained from transformation of GABI_286E07 rpotmp knockout-mutant plants with genetic constructs created in [Tarasenko et al., 2016]. Goal was to determine the effects of RPOTmp knockout/overexpression on global Arabidopsis thaliana gene expression.
Project description:Arabidopsis thaliana (Arabidopsis) encodes five DOUBLE-STRANDED RNA BINDING (DRB) proteins, DRB1 to DRB5, that predominantly act as non-catalytic cofactors for DICER-LIKE (DCL) proteins in the double-stranded RNA (dsRNA) processing stages of small RNA (sRNA) production pathways. In the nucleus, DRB1 is required for microRNA (miRNAs) processing from imperfectly dsRNA precursors by DCL1. Similarly, DRB4 is required by DCL4 for small-interfering RNAs (siRNAs) production from endogenous or exogenous perfectly dsRNA templates. DRB2 has been recently demonstrated to be required for miRNA and siRNA production in developmentally-important tissues of Arabidopsis while the requirement of either DRB3 or DRB5 in sRNA production remains unclear. Here, we analyse in parallel, the contribution of all five DRB protein family members to the global sRNA landscape of Arabidopsis floral tissues. In depth bioinformatic analysis of sRNA sequencing datasets generated from floral tissues of DRB knockout mutant (drb) plant lines, drb1, drb2, drb4, drb12, drb14, drb24, and drb35 and their comparison to the floral sRNA profile of wild-type Arabidopsis, has enabled confident assignment of the requirement of DRB1, DRB2 and DRB4 for the production of specific miRNA and siRNA subclasses in this tissue. Our analyses have additionally identified novel and/or expanded roles for DRB2 in miRNA, trans-acting siRNAs (tasiRNAs) and natural antisense transcript siRNAs (natsiRNAs) production.
Project description:The purpose of the McrBC methylation microarray assay is to determine which regions of a genome are methylated versus those that are unmethylated in a single Arabidopsis thanliana genotype. McrBC is a methylation-sensitive enzyme that restricts DNA only at purine-Cmethyl half sites when separated between 50bp and 3kb. A designated amount of DNA from a particular genotype is sheared to a size range of 1kb-10kb using nebulization. We restrict half of the nebulized DNA with McrBC, and the methylated fraction is then removed from the unmethylated fraction through gel purification of DNA fragments greater than 1kb.* The remaining nebulized DNA is subjected to the same gel purification scheme, but with no McrBC treatment. In a single hybridization, the untreated sample is labeled with Cy5 and the McrBC-treated sample with Cy3. Thus, after labeling and microarray hybridization, the ratio of normalized Cy5 to normalized Cy3 represents the relative methylation at the sequence represented by the spot on the microarray. Dye swap analysis is carried out to take account of experimental variation by repeating the hybridization with identical samples labeled with Cy3 and Cy5, respectively. The two samples in this series are complementary hybridizations in a dye-swap analysis. This is the normalized result of the paired dye swap samples VC121 and VC127. The ANOVA model of Kerr, Martin and Churchill (2000) was used to analyze the data from the dye-swap experiments, with terms included to account for gene, dye-by-gene, treatment-by-gene, and random error terms. These samples were not subjected to hypothesis testing. Therefore, P-values are reported as -1 and no features are flagged as significantly methylated. Keywords: other
Project description:Identification of microRNA expressed in Arabidopsis plants grown at different ambient CO2 concentrations and different ambient temperatures Small RNA sequencing of Arabidopsis wild type ecotype Columbia (Col-0) rossette leaves at bolting stage grown at 430 M-BM-1 50ppm and 810 M-BM-1 50ppm CO2 concentrations and 22 M-BM-1 0.5oC and 28 M-BM-1 0.5oC temperatures
Project description:Wild type Arabidopsis Col-0 plants and mutants depleted of the catalytic subunit of the ribosome-associated N-acetyltransferase A (NatA) complex (amiNAA10) were grown on soil for six weeks under short-day conditions (8h light, light intensity: 100 µE, 21 °C/18 °C temperature (day/night), 50 % humidity). The translatome of both genotypes after 2 hours of light (10 am) was determined by feeding of the trackable Met-analogue azidohomoalanine for 4 hours to leaf disks floated on ½ Hoagland medium containing 50 µM azidohomoalanine in the light.
Project description:Experiment was performed to assess the effect of H1 histones knock-out on DNA methylation under control conditions and a role of H1.3 in maintaining stress-induced menthylation response.