Project description:Non-coding RNA profiling by microarray in an Affymetrix® miRNA Array, version 4.1 (Santa Clara, CA, USA), commercially available by Thermo Fisher Scientific (Waltham, MA) We aimed to profile the expression of the entire population of annotated small non-coding RNAs in the Temporal Cortex (TC) of Alzheimer diseased (AD) individuals compared to age-matched controls.
Project description:To investigate the molecular pathological mechanisms of irritable bowel syndrome with diarrhea (IBS-D) and elucidate the effects of acupuncture on IBS-D colonic mucosa protein abundance in rats, a label-free high-throughput liquid chromatography-tandem mass spectrometry (LC-MS)-based proteomics analysis was used to survey the global changes of colonic mucosa proteins between different groups. A Nano flow Ultimate 3000 HPLC (Dionex Corp, Sunnyvale, CA) coupled online to a Q-Exactive Plus mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA) was used in this project.
Project description:ARPE-19 RPE cells obtained from the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China) were cultured in DMEM medium with 10% FBS (Thermo Fisher Scientific, Waltham, USA) at the incubator with 37 °C, 5% CO2 and 100% humidity. Final concentration of 15 μM curcumin (C7727, Sigma-Aldrich, St. Louis, USA) and 100 μM CoCl4 (10007216, Sinopharm Chemical Reagent, Shanghai, China) were added within serum-free medium for 24 h and 4 h respectively. RNA-seq was performed to investigate the transcriptome alteration.
Project description:To explore the cellular pathways and the molecular functions affected by a novel biogenic amine, (3-HKA), we performed a label free quantitative (LFQ) proteomic analysis of mouse lymphatic endothelial cells (LEC), and primary dendritic cells, treated with IFNgammaplus or minus 3-HKA. Biological triplicates, of total cell lysates, were fractionated by one-dimensional gel electrophoresis (1DEF) and the “in gel” tryptic derived peptides analyzed by nano-LC-ESI-MS/MS on a Q Exactive HF quadrupole orbitrap mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA). LFQ analysis highlighted that 3-HKA downregulated many of the inflammatory pathways, more notably JAK/STAT1 and NFkB, associated with IFNgamma activation.
Project description:Tandem Mass Tag (TMT) technology is an in vitro peptide labeling technique developed by Thermo Fisher Scientific, a peptide labeling technology developed in vitro. The technology uses six (TMTsixplex™ Isobaric Label Reagent Set) or 10 (TMT10plex™ Isobaric Label Reagent Set) isotope labels by labeling peptide specific amino acid sites. In TMT experiments, the relative and absolute amounts of proteins in up to 10 different samples can be compared flexibly by tandem mass spectrometry analysis. In TMT experiments, whole proteins from tissue cells are obtained by protein extraction, and the whole proteins are enzymatically cleaved into peptides by trypsin, and the TMT reagents are used to label all the obtained peptides. The TMT reagent consists of three main components: a reporter group, an equilibrium group and a peptide reaction group.
Project description:We performed RNA-seq to demonstrate that the epithelium of human pluripotent stem cell-derived human intestinal organoids is globally similar to the immature human epithelium and we utilize HIOs to investigate complex host-microbe interactions in this naive epithelium. RNA was isolated using the mirVana RNA isolation kit and following the \Total RNA\ isolation protocol (Thermo-Fisher Scientific, Waltham MA). RNA library preparation and RNA-sequencing (single-end, 50 bp read length) were performed by the University of Michigan DNA Sequencing Core using the Illumina Hi-Seq 2500 platform. Transcriptional quantitation analysis was conducted using 64-bit Debian Linux stable version 7.10 (\Wheezy\). Pseudoalignment of RNA-seq data was computed using kallisto v0.43.0 and differential expression of pseudoaligned sequences was calculated using the R package DEseq2.
Project description:ER-positive tumor data was derived by RNA-sequencing of formalin-fixed paraffin-embedded specimens (FFPE) collected from 44 women diagnosed with ER+ and progesterone receptor-positive but HER2 negative (ER+/PR+/HER2-) breast cancer at the Karmanos Cancer Institute. According to a pathologist review, all tumor specimens were invasive cancer with 70% or greater cancer cell content. Patients had not received neoadjuvant cancer therapy, nor were they treated for diabetes with systemic drugs (e.g., metformin) before surgical resec- tion. Following manufacturer instructions, the Recoverall Nucleic Acid Isolation Kit for FFPE Tissue (Ambion, Austin, TX) was used to isolate RNA. Quantitation and fragmentation analysis were performed using the NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA) and Bioanalyzer 2100 and RNA 6000 Nano kit (Agilent Technologies, Santa Clara, CA). A Qubit fluorometric assay (Thermo Fisher Scientific) was used to quantitate and equalize dsDNA prepared using the Truseq RNA Access library preparation kit (Illu- mina, San Diego, CA). Libraries were randomized, indexed, multiplexed, and paired end sequenced (200 cycles total) on an Illumina Nextseq500. Bioinformatic quality control was done using the FastQC software (v0.11.4), adapter and low-quality reads were removed using Trimmomatic v0.36. FPKM normalization and aligning were done using TopHat v2.1.1.
