Project description:In this article we report the identification of T-DNA (transfer DNA) insertion sites within two different gene regions in the genome of an Arabidopsis mutant line, SALK_084889. The T-DNA positions are in the 3' UTR (untranslated region) of DREB2A (Dehydration-responsive element-binding protein 2A) (AT5G05410) and promoter of LOX1 (Lipoxygenase 1) (AT1G55020) as determined by DNA-PCR and sanger sequencing. The expression levels of DREB2A and LOX1 were also analyzed using quantitative realtime PCR (qPCR) in SALK_084889 and wild type Arabidopsis (Col, Columbia). Further, the comparison of drought and heat tolerance between Col and SALK_084889 were conducted by stress treatments. The present data indicate that in SALK_084889, the expression of DREB2A is not downregulated under normal growth conditions but can be affected only in roots under drought treatment, while LOX1 is significantly downregulated in both roots and shoots under all tested conditions. These data are original and have not been published elsewhere.
Project description:In this article we report the identification of a homozygous lethal T-DNA (transfer DNA) line within the coding region of the At1G05290 gene in the genome of Arabidopsis thaliana (Arabidopsis) line, SALK_063500. The T-DNA insertion is found within exon one of the AT1G05290 gene, however a homozygous T-DNA allele is unattainable. In the heterozygous T-DNA allele the expression levels of AT1G05290 were compared to wild type Arabidopsis (Col-0, Columbia). Further analyses revealed an aberrant silique phenotype found in the heterozygous SALK_063500 plants that is attributed to the reduced rate of pollen tube germination. These data are original and have not been published elsewhere.
Project description:Some aquaporins do not show a pronounced function as water diffusion facilitators but act as small molecule transport facilitators for substances such as urea, glycerol, boron or gases such as CO2 . Transcriptome analysis provided distinguishable, specific profiles for water stress or for conditions of increased or decreased CO2 concentrations We used Affymetrix microarray analysis to elucidate the function of the aquaporin AtPIP1;2 gene
Project description:BackgroundThe three-dimensional (3D) organization of chromosomes is linked to epigenetic regulation and transcriptional activity. However, only few functional features of 3D chromatin architecture have been described to date. The KNOT is a 3D chromatin structure in Arabidopsis, comprising 10 interacting genomic regions termed KNOT ENGAGED ELEMENTs (KEEs). KEEs are enriched in transposable elements and associated small RNAs, suggesting a function in transposon biology.ResultsHere, we report the KNOT's involvement in regulating invasive DNA elements. Transgenes can specifically interact with the KNOT, leading to perturbations of 3D nuclear organization, which correlates with the transgene's expression: high KNOT interaction frequencies are associated with transgene silencing. KNOT-linked silencing (KLS) cannot readily be connected to canonical silencing mechanisms, such as RNA-directed DNA methylation and post-transcriptional gene silencing, as both cytosine methylation and small RNA abundance do not correlate with KLS. Furthermore, KLS exhibits paramutation-like behavior, as silenced transgenes can lead to the silencing of active transgenes in trans.ConclusionTransgene silencing can be connected to a specific feature of Arabidopsis 3D nuclear organization, namely the KNOT. KLS likely acts either independent of or prior to canonical silencing mechanisms, such that its characterization not only contributes to our understanding of chromosome folding but also provides valuable insights into how genomes are defended against invasive DNA elements.
Project description:Some aquaporins do not show a pronounced function as water diffusion facilitators but act as small molecule transport facilitators for substances such as urea, glycerol, boron or gases such as CO2 . Transcriptome analysis provided distinguishable, specific profiles for water stress or for conditions of increased or decreased CO2 concentrations We used Affymetrix microarray analysis to elucidate the function of the aquaporin AtPIP1;2 gene A T-DNA insertion line atpip1;2-1 was used for RNA extraction and further microarray analysis based on the Affymetrix platform. The transcriptome of the atpip1;2-1 line was compared to the wild type (N-60000).
Project description:Despite the growing volume of experimentally validated knowledge about the subcellular localization of plant proteins, a well performing in silico prediction tool is still a necessity. Existing tools, which employ information derived from protein sequence alone, offer limited accuracy and/or rely on full sequence availability. We explored whether gene expression profiling data can be harnessed to enhance prediction performance. To achieve this, we trained several support vector machines to predict the subcellular localization of Arabidopsis thaliana proteins using sequence derived information, expression behavior, or a combination of these data and compared their predictive performance through a cross-validation test. We show that gene expression carries information about the subcellular localization not available in sequence information, yielding dramatic benefits for plastid localization prediction, and some notable improvements for other compartments such as the mitochondrion, the Golgi, and the plasma membrane. Based on these results, we constructed a novel subcellular localization prediction engine, SLocX, combining gene expression profiling data with protein sequence-based information. We then validated the results of this engine using an independent test set of annotated proteins and a transient expression of GFP fusion proteins. Here, we present the prediction framework and a website of predicted localizations for Arabidopsis. The relatively good accuracy of our prediction engine, even in cases where only partial protein sequence is available (e.g., in sequences lacking the N-terminal region), offers a promising opportunity for similar application to non-sequenced or poorly annotated plant species. Although the prediction scope of our method is currently limited by the availability of expression information on the ATH1 array, we believe that the advances in measuring gene expression technology will make our method applicable for all Arabidopsis proteins.
Project description:Two minichromosomes (alpha and delta) in addition to two other aberrant chromosomes (beta and gamma) were found in a transgenic Arabidopsis plant produced by an in planta vacuum infiltration technique. The minichromosomes were successfully separated by successive crossing and selfing and added to wild-type Columbia (Col-0) as a supernumerary chromosome. FISH indicated that both of the two minichromosomes originated from the short arm of chromosome 2. The mini alpha chromosome contained the whole short-arm 2S and a truncated centromere (180-bp repeat cluster), whereas mini delta lacked the terminal region including telomere repeats. Pachytene FISH clearly revealed that mini delta comprised a ring chromosome carrying two copies of the region from the 180-bp repeat cluster to BAC-F3C11. Both of the 180-bp clusters (each approximately 500 kb in length) were thought to possess normal centromere functions because the centromere-specific histone H3 variant (HTR12) was detected on both clusters. Notwithstanding this dicentric and ring form, mini delta was stably transmitted to the next generations, perhaps because of its compact size (<4 Mb). Chromosome beta also comprised a dicentric-like structure, with one of the two 180-bp repeat sites derived from chromosome 1 and the other from chromosome 2. However, the latter was quite small and failed to bind HTR12. The data obtained in this study indicated that 500 kb of the 180-bp array of the chromosome 2 centromere, from the edge of the 180-bp array on the short-arm side, is sufficient to form a functional domain.
Project description:Ordered collections of Arabidopsis thaliana lines containing mapped T-DNA insertions have become an important resource for plant scientists performing genetic studies. Previous reports have indicated that T-DNA insertion lines can have chromosomal translocations associated with the T-DNA insertion site, but the prevalence of these rearrangements has not been well documented. To determine the frequency with which translocations are present in a widely-used collection of T-DNA insertion lines, we analyzed 64 independent lines from the Salk T-DNA mutant collection. Chromosomal translocations were detected in 12 of the 64 lines surveyed (19%). Two assays were used to screen the T-DNA lines for translocations: pollen viability and genome-wide genetic mapping. Although the measurement of pollen viability is an indirect screen for the presence of a translocation, all 11 of the T-DNA lines showing an abnormal pollen phenotype were found to contain a translocation when analyzed using genetic mapping. A normal pollen phenotype does not, however, guarantee the absence of a translocation. We observed one T-DNA line with normal pollen that nevertheless had a translocation based on genetic mapping results. One additional phenomenon that we observed through our genetic mapping experiments was that the T-DNA junctions on the 5'- and 3'-sides of a targeted gene can genetically separate from each other in some cases. Two of the lines in our survey displayed this 'T-DNA borders separate' phenomenon. Experimental procedures for efficiently screening T-DNA lines for the presence of chromosomal abnormalities are presented and discussed.