Project description:We report the application of ChIP-Seq technology for analyzing the DNA binding sites of SOD1 in the nucleus of HeLa cells. By obtaining a plenty of sequence from chromatin immunoprecipitated DNA, we generated genome-wide DNA binding sites of SOD1. After sequencing of ChIP samples, 42,737,195, 49,950,032, and 38,825,768 clean reads for control group, H2O2 treated group and LD100 (a specific inhibitor of SOD1) treated group were obtained through trimming the raw reads. We find that SOD1 occupies DNA sites with distinct sequence preference in the nucleus. The treatment with either H2O2 or LD100 was found to decrease the strength of SOD1 binding to DNA, indicating that the H2O2 exposure- or SOD1 inhibition-mediated redox dyshomeostasis may result in decreased genes that are reasonably regulated through alteration of SOD1 structures compared to control.
Project description:Contaminated aquifer (Dusseldorf-Flinger, Germany) templates extracted from 5 sediment depths ranging between 6.4 and 8.4 m below ground and over 3 years of sampling were amplified for amplicon pyrosequencing using the primers Ba27f (5’-aga gtt tga tcm tgg ctc ag-3’) and Ba519r (5’- tat tac cgc ggc kgc tg-3’), extended as amplicon fusion primers with respective primer A or B adapters, key sequence and multiplex identifiers (MID) as recommended by 454/Roche. Amplicons were purified and pooled as specified by the manufacturer. Emulsion PCR (emPCR), purification of DNA-enriched beads and sequencing run were performed following protocols and using a 2nd generation pyrosequencer (454 GS FLX Titanium, Roche) as recommended by the developer. Quality filtering of the pyrosequencing reads was performed using the automatic amplicon pipeline of the GS Run Processor (Roche), with a slight modification concerning the valley filter (vfScanAllFlows false instead of TiOnly) to extract the sequences. Demultiplexed raw reads were furhter trimmed for quality and lenght (>250 bp).
Project description:Purpose: The goal of this study is to compare endothelial small RNA transcriptome to identify the target of OASL under basal or stimulated conditions by utilizing miRNA-seq. Methods: Endothelial miRNA profilies of siCTL or siOASL transfected HUVECs were generated by illumina sequencing method, in duplicate. After sequencing, the raw sequence reads are filtered based on quality. The adapter sequences are also trimmed off the raw sequence reads. rRNA removed reads are sequentially aligned to reference genome (GRCh38) and miRNA prediction is performed by miRDeep2. Results: We identified known miRNA in species (miRDeep2) in the HUVECs transfected with siCTL or siOASL. The expression profile of mature miRNA is used to analyze differentially expressed miRNA(DE miRNA). Conclusions: Our study represents the first analysis of endothelial miRNA profiles affected by OASL knockdown with biologic replicates.
Project description:Chaetomium globosum is a model of conditional pathogens abundant on a wide variety of substrates in soil, water, and atmosphere environments. Homothallic C. globosum produces hairy perithecia bearing meiotic ascospores that are resistant to harsh conditions for dispersal, and asexual reproduction of conidia has never been observed. RNAs were samples from nine distinct morphological stages during the nearly synchronic perithecial development for C. globosum. Unlike the heterothallic Neurospora crassa, the mating type gene mat a-1 showed comparatively lower expression changes but highly coordinate with expression regulation of mat A-1 in C. globosum. Key regulators, including orthologs of N. crassa sub-1, sub-1 dependent gene NCU00309, and asl-1, in the initiation of sexual development in response to light stimuli, showed similar regulation dynamics between C. globosum and N. crassa. Knockout phenotyping directed by the comparative analysis of transcriptomics between C. globosum and its’ closely related Neurospora crassa also suggested some genes that are critical for perithecial development. Among 24 secondary metabolism clusters composed more than 3 genes in C. globosum, 11 showed highly coordinated expression across the perithecial development, and dramatically up-regulation was recorded for all 12 genes in the cochliodones biosynthesis cluster. Up-regulation of chaetoglocin and aureonitol biosynthesis clusters was found to be associated with disturbance in early sexual development and with ascospore maturation. Similar to pathogenic Fusarium graminearum, C. globosum showed coordinately up-regulated expression of homologs of histidine kinases in hyperosmotic response pathways, consist with their ecology adapting to high humidity.
Project description:A cDNA library was constructed by Novogene (CA, USA) using a Small RNA Sample Pre Kit, and Illumina sequencing was conducted according to company workflow, using 20 million reads. Raw data were filtered for quality as determined by reads with a quality score > 5, reads containing N < 10%, no 5' primer contaminants, and reads with a 3' primer and insert tag. The 3' primer sequence was trimmed and reads with a poly A/T/G/C were removed