HEK293 cells 100 cell RNAseq profiling on Ion Proton
ABSTRACT: Five libraries from 100 HEK293 cells each were prepared using a Smartseq based custom library preparation approach with unique molecular identifiers. One batch of 2 replicates (A) and one batch of 3 replicates (B) were prepared from different cell cultures. Libraries were sequenced on an Ion Proton HEK293 cell (100 cells) 5' selective RNAseq profiling, N4H4 unique molecular identifiers, 2 replicates (A) and 3 replicates (B)
Project description:Three libraries from 100 HEK293 cells each were prepared using a Smartseq based custom library preparation approach with unique molecular identifiers. Libraries were sequenced on a Illumina NextSeq 500 HEK293 cell (100 cells) 5' selective RNAseq profiling, N4H4 unique molecular identifiers, 3 replicates
Project description:We have applied a recently developed, highly accurate and sensitive single-cell RNA-seq method (STRT/C1) to perform a molecular census of two regions of the mouse cerebral cortex: the somatosensory cortex and hippocampus CA1. We isolated cells fresh from somatosensory cortex (S1) and hippocampus CA1 area of juvenile (P22 - P32) CD1 mice, 33 males and 34 females. Cells were collected without selection, except that 116 cells were obtained by FACS from 5HT3a-BACEGFP transgenic mice. A total of 76 Fluidigm C1 runs were performed, each attempting 96 cell captures and resulting in 3005 high-quality single-cell cDNAs, containing Unique Molecular Identifiers allowing counting of individual mRNA molecules, even after PCR amplification.
Project description:Purpose: We applied cDNA molecule counting using unique molecular identifiers combined with high-throughput sequencing to study the transcriptome of individual mouse embryonic stem cells, with spike-in controls to monitor technical performance. We further examined transcriptional noise in the embryonic stem cells. One 96-well plate of single-stranded cDNA libraries generated from 96 single R1 mouse embryonic stem cells sequenced on two lanes, and one 96-well plate of the same libraries further amplified by 9 PCR cycles sequenced on one lane.
Project description:Single-cell RNA-seq from CD4+ T lymphocytes from uninfected steady-state mouse, two mice with Salmonella typhimurium infection at day 14 and one mouse at day 49 post-infection. Used to demonstrate application of reconstruction and analysis of T cell receptor sequences from single-cell RNA-seq.
Project description:Understanding cell type identity in complex tissues or organisms requires integration of each cell's expression profile with its spatial location within the tissue under study. We developed a high-throughput method that combines in vitro single-cell RNA-sequencing with a gene expression atlas to map single cells back to their location within the tissue of interest. We used the developing brain of a marine annelid, Platynereis dumerilii that is an important model system for studying bilaterian brain evolution, to benchmark our approach. To generate the single-cell mRNA-sequencing data, P. dumerilii larval brains were dissociated, followed by cell capture, cDNA synthesis and amplification on the C1 Single-Cell Auto Prep IFC for 10-17 um cells (Fluidigm). Sequencing libraries were produced using Nexera XT DNA kit (Illumina). In total, we sequenced 213 samples, of which 129 correspond to single, alive cells (as judged by visual inspection of the captured cells) with the remainder consisting of a variety of single dead cells (n=18), wells containing extracellular matrix contaminants (n=8) or multiple cells (n=17), as well as a negative controls where no cells were observed (n=41). For this dataset, we achieved ~90% success rate for the spatial mapping of the single-cell RNA-seq data to P. dumerilii brain atlas. NOTE: 72 additional samples were added on 13th December 2014.
Project description:We analysed a well-established fraction of mouse subcutaneous adipose-derived SVF cells that is generally considered to harbour adipose stem and progenitor cells (ASPCs). To study the molecular characteristics and the subpopulation structure of ASPCs, we performed three replicate scRNA-seq experiments. We collected Lin- (CD31- CD45- TER119-) CD29+ CD34+ SCA1+ cells from the mouse subcutaneous SVF of transgenic mice, in which red fluorescent protein (RFP) is induced in Dlk1-expressing cells upon feeding with tamoxifen. While CD29, CD34, and SCA1 are generally expected to enrich for stem cells, DLK1 has previously been suggested to specifically mark pre-adipocytes. The submission includes the raw data of all sequenced cells, while we only processed 208 high quality single cells. RFP status is indicated in the processed files.
Project description:In this study, we assess technical differences between commonly used single-cell RNA-Sequencing (scRNA-Seq) methods. We perform scRNA-seq on a homogenous population of mouse embryonic stem cells along with two kinds of control spike-in molecules to assess sensitivity and accuracy of these specific methods. In this dataset, we perform Smart-Seq2 method on Fluidigm C1 system and generate single-cell libraries using Nextera XT kit
Project description:In this study, we assess technical differences between commonly used single-cell RNA-Sequencing (scRNA-Seq) methods. We perform scRNA-seq on a homogenous population of mouse embryonic stem cells along with two kinds of control spike-in molecules to assess sensitivity and accuracy of these specific methods. In this dataset, we perform SMARTer method on Fluidigm C1 system and generate single-cell libraries using Nextera XT kit