Project description:Analysis of leaves of wild-type and rice COI mutants treated with methyl jasmonate (MeJA). Results provide the role of rice COI on response to jasmonic acid.
Project description:Aim: To improve risk stratification in patients with stable coronary artery disease (CAD), we aimed to identify genes in monocytes predictive of new ischemic events in patients with CAD and determine to what extent expression of these transcripts resembles expression in acute myocardial infarction (AMI). Results: COX10 and ZNF484 distinguished between AMI and the whole group of stable CAD patients with an accuracy of 90%. COX10 and ZNF484 together with MT-COI and WNK1 distinguished AMI patients from stable CAD patients with and without a new event with a sensitivity of 89% and a specificity of 98%. MT-COI and COX10 increased the accuracy for separating stable CAD patients with and without a new coronary event from 68 to 80% in addition to age, gender, BMI, diabetes, lipids, blood pressure and hs-CRP. Interestingly, expression of MT-COI, COX10 and WNK1 (but not ZNF484) in PBMCs paired with that in monocytes; COX10 in whole blood was similar to that in monocytes. Conclusions: This work showed that COX10 and ZNF484, eventually combined with MT-COI and WNK1 have the potential to accurately discriminate between AMI and stable CAD patients, and may improve the risk assessment of stable CAD patients.
Project description:Coilin is a scaffold protein essential for the structural integrity of Cajal Bodies, which are non-membranous nuclear organelles that are thought to facilitate assembly and maturation of nuclear RNPs, including spliceosomal snRNPs. To investigate further coilin’s functions in plant cells, and to identify proteins that may functionally interact with coilin, we performed a genetic suppressor screen in Arabidopsis thaliana using a coilin (coi) mutant displaying altered splicing of a GFP pre-mRNA. The modified splicing pattern results in a ‘hyper-GFP’ phenotype in young coi seedlings relative to the intermediate level of GFP in wild-type seedlings. Additionally, in newly emerging leaves of older coi seedlings, the GFP gene frequently undergoes abrupt siRNA-associated posttranscriptional gene silencing that persists during growth. In the suppressor screen, we searched for mutations that subdue one or both of these GFP phenotypes and identified several understudied factors in plants: WRAP53, a putative Cajal body protein; SMU2, a predicted splicing-related factor; and ZC3HC1, an uncharacterized zinc finger protein. All three mutations return the hyper-GFP phenotype of the coi mutant to approximately the intermediate wild-type level. The zc3hc1 mutations in particular induce premature and more extensive posttranscriptional gene silencing similar to mutations in SOP1 and DCL4, which are known modifiers of posttranscriptional gene silencing. Candidate coilin-interacting proteins identified by immunoprecipitation-mass spectrometry include many splicing-related factors, nucleolar proteins, and mRNA export factors. Our results demonstrate the usefulness of the coi mutant to identify new modifiers of alternative splicing and posttranscriptional gene silencing, and suggest diverse roles for coilin in plant cells.
Project description:We determined circRNA abundance in fly Heads and S2 cells by generating and analyzing high-throughput RNA-sequencing libraries prepared from rRNA-depleted RNA. In order to determine whether the observed sequencing reads are due to bona fide circRNAs, we pre-treated the RNA with RNAse-R before the rRNA-depletion procedure. Indeed, most of the identified circRNAs were more enriched in comparison to the canonical mRNA isoforms following the RNAse-R treatment. We compare circRNA levels in wt (Canton S) flies with flies carrying the C4 ("slow polymerase") mutation. 4 samples of Drosophila Canton S and 4 samples of flies carrying the C4 ("slow polymerase") mutation. For each sample, one library was prepared from RNA after RNaseR treatment and the second from RNA with without treatment (mock). RNA library from one Canton Sample was used for stranded libray preprepation.
Project description:The importance of single-cell level data is increasingly appreciated, and significant advances in this direction have been made in recent years. Common to these technologies is the need to physically segregate individual cells into containers, such as wells or chambers of a micro-fluidics chip. High-throughput Single-Cell Labeling (Hi-SCL) in drops is a novel method that uses drop-based libraries of oligonucleotide barcodes to index individual cells in a population. The use of drops as containers, and a microfluidics platform to manipulate them en-masse, yields a highly scalable methodological framework. Once tagged, labeled molecules from different cells may be mixed without losing the cell-of-origin information. Here we demonstrate an application of the method for generating RNA-sequencing data for multiple individual cells within a population. Barcoded oligonucleotides are used to prime cDNA synthesis within drops. Barcoded cDNAs are then combined and subjected to second generation sequencing. The data are deconvoluted based on the barcodes, yielding single-cell mRNA expression data. In a proof-of-concept set of experiments we show that this method yields data comparable to other existing methods, but with unique potential for assaying very large numbers of cells. In this experiment we mixed 2 cell types (mES mEF) and then using single cell novel approach we could be able to find each cell (using its barcode) and assign it to mES of mEF and to produce mES and mEF aggregate bam files (converted to bed for GEO submission). 1152_RNA_RTprimers_Barcodes.txt: A list of all 1152 barcodes sequenced for Read2 fastq files.
Project description:We describe a novel workflow named Barcode Assembly foR Targeted Sequencing, which is a highly sensitive, quantitative, and inexpensive technique for targeted sequencing of transcript cohorts (rBART-Seq) or genomic regions (gBART-Seq) from thousands of bulk samples or single cells in parallel. Multiplexing is based on a simple method that produces extensive matrices of diverse DNA barcodes attached to invariant primer sets, for generating amplicons with dual indices. Here, we used the gBART-Seq for genetic screening of breast cancer patients and identified BRCA mutations with very high precision.