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Single Cell Molecular Profiling

Single Cell RNAseq

Single cell transcription profiling provides the opportunity to stratify complex cell types from heterogeneous tissue, to identify rare cell types, and to characterize subpopulations from seemingly homogeneous populations of cells. Given the ability of the technique to identify subtle expression changes at a single cell level, the approach is ideal for developing normative expression in atlas-type studies or assessing the impact that disease or treatment states may have on tissues and cells. The Molecular Genomics Core has been carrying out scRNAseq since 2016 and in that time has run over 300 samples from over 30 investigators across 9 different species, giving us confidence in the data we generate.

Coming soon: Targeted Gene Expression
Following the wide-cast net of 10x Genomics’ single cell gene expression assay, explore the most relevant cell types and biomarkers with targeted gene panels. Scale up gene expression experiments without the sequencing costs of whole transcriptome. Choose a panel with comprehensive content (cancer, immunology, neuroscience, and gene pathways) or create custom panels.

 

Single Nuclei RNAseq (snRNA-seq)

Optimally, single cell RNA sequencing data is generated from freshly dissociated cells with high viability. However, in some circumstances it is challenging to collect fresh biospecimens, the tissue may be a legacy sample from frozen storage, or tissue dissociation may cause lysing of specifically labile cell types. To circumvent these problems, and with development support from the Duke SoM, the MGC has optimized nuclei isolation protocols to generate snRNA-Seq data. We have applied our new protocol to a two-year-old frozen brain tumor sample to assess the use of legacy samples, and fresh mouse brain tissue to capture sensitive neuronal expression profiles, using the 10xGenomics Chromium platform.

For more information, visit 10xGenomics.com.

Sample Requirement Information Sheet Cell Prep Guide

 

Single Cell Assay for Transposase Accessible Chromatin (scATAC-seq)

The tightly packed eukaryotic genome relies on the highly regulated alteration of chromatin structure to facilitate the accessibility of genomic regions for gene expression. These chromatin alterations within the genome can reveal a great deal about the state of the cell. Assay for Transposase-Accessible Chromatin (ATAC-seq) coupled with next-generation sequencing is a technique to identify accessible chromatin regions. The 10x Genomics’ Chromium Single Cell ATAC Sequencing Solution (scATAC-Seq) is capable of identifying rare and important cellular subtypes that cannot be acquired in amounts sufficient for genome-wide chromatin analyses. With as little as a few hundred cells, scATAC-Seq allows profiling of chromatin structure at the single cell level, providing an unprecedented view of the heterogeneity of chromatin regulation.

Coming soon: Single Cell ATAC + Gene Expression
Unite transcriptional and chromatin landscape by combining gene expression and ATAC-seq data from the same cell. Characterize direct linkages between genes and regulatory elements across tens of thousands of cells.

For more information, visit 10xGenomics.com.

 

Single Cell Immune Profiling

The ability to profile immune receptor clonotypes confers the ability to predict the success of stem cell therapy, establish the impact of antigenic exposure, and characterize the properties of tumor infiltrating lymphocytes, amongst other applications. Previous approaches have been limited to heavy chain profiling of specificity conferring regions of the genome, however the 10x Genomics Single Cell V(D)J application offers a more comprehensive solution for measuring immune repertoire information and gene expression from the same single cell. Importantly, paired T-cell receptor (TCR) or B-cell immunoglobulin (Ig) receptor sequences are profiled from 100-10,000 individual cells per sample.

For more information, visit 10xGenomics.com.

 

Single Cell Feature Barcoding Technology

Single cell expression profiling via scRNA-Seq and snRNA-Seq provides an invaluable snapshot of cell state, but does not predict the translational outcome of mRNA expression. To do this, 10x Genomics have adopted Feature Barcoding technology that uses their microfluidic platform to assess concomitantly single cell gene and protein expression. A pool of ~3,500,000 10x Barcodes separately index each cell’s transcriptome and surface proteins by partitioning between 500 – 10,000 cells using the platforms nanoliter-scale gel bead technology. Protein specific antibodies with custom ligated adapters are bound to the target cells, cells are partitioned into specific GEMs, and proteins and transcripts are assigned to a specific cell via barcodes, and then proteins and mRNA are quantitatively measured using unique molecular indices. The MGC is currently using development funds from the Duke Cancer Institute to test the approach and concomitantly profile mRNA and protein in several different tissue types.

Future Development for Single Cell Feature Barcoding

Single Cell Gene Expression and Crispr Screening

Highlights

  • Simultaneously assess perturbation phenotypes and gene expression from the same cell
  • Enable high throughput and high resolution functional genetic screens in hundreds to tens of thousands of cells simultaneously
  • Determine comprehensive gene expression phenotypes for individual perturbations
  • Customize your CRISPR pools with dozens, to hundreds, to thousands of gRNAs
  • Implement an improved and novel methodology over published methods by directly capturing and sequencing gRNAs, eliminating the need for a proxy barcode

For more information, visit 10xGenomics.com.

 

Cell Multiplexing

Coming soon: Cell Multiplexing
Leverage CellPlex for sample multiplexing to enable flexible experimental design with greater throughput for your single cell gene expression and immune profiling experiments. Combine samples or profile more cells across diverse conditions, including varied primary samples, time-course studies, dose-response titrations, and more, in a single run. Combine multiple samples in one multiplexed library to save on library generation costs.

Intracellular Protein Detection

Coming soon: Intracellular Protein Detection
Detect gene expression and intracellular proteins from the same cell with Feature Barcode technology. Profile key transcription factors at both the protein and transcript level. Deepen your understanding of the gene-protein regulatory networks that govern development and disease progression. Assess post-translational modifications of target proteins.