RNA-seq in Arabidopsis thaliana Col and hcr2 mutant
Ontology highlight
ABSTRACT: RNA-prep of Col and hcr2 mutant from seedling and bud sample. RNA extracted by Trizol and cDNA was synthesized by using the ScriptSeq v2 RNA-seq Library Preparation Kit (SSV21124, Epicentre). ScriptSeq Index PCR Primers (RSBC10948, Epicentre) were used for library construction.
Project description:RNA-prep of Col and hcr3 mutant from seedling or bud sample. RNA was extracted using the RNeasy plant (Qiagen, 74904) and sent to BGI Hongkong for RNA-seq library construction and sequencing. Briefly, mRNA enriched by using oligo dT beads were fragmented and used for cDNA synthesis. dUTP method was used for cDNA synthesis to make the RNA-seq library strand-specific. The constructed library was amplified to make DNA nanoball (DNB) and sequenced on DNBSEQ platform.
Project description:RNA-prep of j3-1, j3-3, j2-2, hcr3 mutant from seedling or bud sample. RNA was extracted using the RNeasy plant (Qiagen, 74904) and sent to BGI Hongkong for RNA-seq library construction and sequencing. Briefly, mRNA enriched by using oligo dT beads were fragmented and used for cDNA synthesis. dUTP method was used for cDNA synthesis to make the RNA-seq library strand-specific. The constructed library was amplified to make DNA nanoball (DNB) and sequenced on DNBSEQ platform.
Project description:Col and hcr2 mutant from seedling and bud sample were harvested. Nuclei extracted using Honda buffer. Cross-linked chromatin was fragmented with mirococcal nuclease (MNase, NEB M0247S). After fragmentation, fragmented chromatin was incubated with H3K4me3 antibody (AbCam ab8580). TruSeq Prep Kit v2 (Illumina) were used for library construction.
Project description:DNA-prep of Col and hcr2 mutant from seedling and bud sample. DNA was extracted using DNeasy Plant Mini Kit (Qiagen 69104, USA). Library construction and bisulfite conversion was performed using EZ DNA Methylation-Gold kit (ZYMO). DNA libraries are sequenced on the DNBseq platform (BGI, Hong Cong).
Project description:ATAC-seq of Col and hcr2 mutant from seedling and bud sample. Tagmentation was performed using the Tag DNA enzyme & buffer kit (Illumina, 20034210). Nextera DNA CD Index primers were used for library construction. The indexed libraries were subjected to paired-end 50-bp sequencing using an Illumina HiSeqX instrument (Microgen, Korea).
Project description:To investigate the impact of disruption of the non-CG DNA methylation/H3K9me2 pathway upon transcription in Arabidopsis, we performed RNA-seq using meiotic-stage floral buds from wild type (Col-0) and kyp/suvh4 suvh5 suvh6 mutant plants. This enabled identification of differentially expressed genes and transposable elements (TEs). TEs that were up-regulated in kyp/suvh4 suvh5 suvh6 relative to wild type were evaluated for over-representation of elements within each TE family.
Project description:Advantages of RNA-Seq over array based platforms are quantitative gene expression and discovery of expressed single nucleotide variants (eSNVs) and fusion transcripts from a single platform, but the sensitivity for each of these characteristics is unknown. We measured gene expression in a set of manually degraded RNAs, nine pairs of matched fresh-frozen, and FFPE RNA isolated from breast tumor with the hybridization based, NanoString nCounter, (226 gene panel) and with whole transcriptome RNA-Seq using RiboZeroGold ScriptSeq V2 library preparation kits. We performed correlation analyses of gene expression between samples and across platforms. We then specifically assessed whole transcriptome expression of lincRNA and discovery of eSNVs and fusion transcripts in the FFPE RNA-Seq data. For gene expression in the manually degraded samples, we observed Pearson correlation of >0.94 and >0.80 with NanoString and ScriptSeq protocols respectively. Gene expression data for matched fresh-frozen and FFPE samples yielded mean Pearson correlations of 0.874 and 0.783 for NanoString (226 genes) and ScriptSeq whole transcriptome protocols respectively. Specifically for lincRNAs, we observed superb Pearson correlation (0.988) between matched fresh-frozen and FFPE pairs. FFPE samples across NanoString and RNA-Seq platforms gave a mean Pearson correlation of 0.838. In FFPE libraries, we detected 53.4% of high confidence SNVs and 24% of high confidence fusion transcripts. Sensitivity of fusion transcript detection was not overcome by an increase in depth of sequencing up to 3-fold (increase from ~56 to ~159 million reads). Both NanoString and ScriptSeq RNA-Seq technologies yield reliable gene expression data for degraded and FFPE material. The high degree of correlation between NanoString and RNA-Seq platforms suggests discovery based whole transciptome studies from FFPE material will produce reliable expression data. The RiboZeroGold ScriptSeq protocol performed particularly well for lincRNA expression from FFPE libraries but detection of eSNV and fusion transcripts was less sensitive. We performed RNASeq on RNA from nine matched pairs of fresh-frozen and FFPE tissues from breast cancer patients. The goal was to test the RiboZeroGold ScriptSeq complete low input library preparation kit for degraded RNA samples.
Project description:We sequenced messenger RNA isolated from seeding, root, and floral bud tissue for 2 MAGIC founder accessions of Arabidopsis thaliana. The resulting data was used for validation of gene models, and provides a resource to assess tissue-specific expression. Examination of RNA expression across tissues (seedling, root, floral bud) for 2 accessions (Col-0, Can-0).
Project description:Three-day metatranscriptome of surface gravel plain soils from the Central Namib Desert. Samples were collected at four times (6:00, 12:00, 18:00 and 24:00h) on each day (n=12). rRNA-depleted RNA was used to construct stranded libraries with the ScriptSeq v2 complete kit (Epicentre) adding unique barcodes in TruSeq adapters (ScriptSeq Index PCR primers, set 1, Epicentre). Libraries were single-end sequenced in a NextSeq 500 v2 sequencer, with read length of 75bp.
Project description:To investigate the influence of heterochromatin on transposon transcription, we performed sequencing of RNA in wild type plant and met1 mutant which is defective in maintenance of CG context DNA methylation. The met1 mutant showed de-repression of centromeric transposons including Gypsy and EnSpm, which is coincident with increased meiotic DSBs and decreased nucleosome occupancy.