10x Genomics single cell library preps

The Core offers single cell sequencing using 10x Genomics’ Chromium Controller and library preparation kits. A variety of kit options are available – please see individual descriptions below.

A consultation is required prior to submitting an order. Please contact Dr. Katia Sol-Church (ks5uq@virginia.edu) to set up a meeting time.

After you have discussed your project, please place an order in iLab using our ’10xGenomics library prep and sequencing NGS’ project request. A form will guide you through the order placement process. If you have any questions about placing your order, please contact either Dr. Katia Sol-Church (ks5uq@virginia.edu) or Alyson Prorock (ajp7x@virginia.edu) for assistance.

10x Genomics 5′ Next GEM Gene Expression Profiling library prep

The Chromium Single Cell 5′ GEX library prep technology offers a comprehensive, scalable approach to profiling gene expression from up to 10,000 single cells per library, allowing for deep profiling of cell types within complex cell populations. Full-length (5’ UTR to constant region) transcripts from 100-10,000 individual cells per sample can be assessed by partitioning cells into nanoliter-scale Gel Beads-in-emulsion (GEMs) using the 10x Genomics Chromium Controller, where all generated cDNA share a common 10x Barcode. Libraries are generated and sequenced and 10x Barcodes are used to associate individual reads back to the individual cell partitions.

Download the 10x Genomics user guide here: 5′ Gene Expression Profiling user guide

Uses Chromium Next GEM Single Cell 5′ Library and Gel Bead Kit v1.1 (10x Genomics, 1000165 or 1000167) and
Chromium Next GEM Chip G Single Cell Kit (10x Genomics, 1000120/1000127)

10x Genomics VDJ immune profiling (B cell or T cell) library prep

The Single Cell V(D)J protocols offer comprehensive, scalable solutions for measuring immune repertoire information and gene expression from the same cell. Profile full-length (5’ UTR to constant region), paired T-cell receptor (TCR), or B-cell immunoglobulin (Ig) transcripts from 100-10,000 individual cells per sample. A pool of ~750,000 barcodes are sampled separately to index each cell’s transcriptome. It does so by partitioning thousands of cells into nanoliter-scale Gel Beads-in-emulsion (GEMs), where all generated cDNA share a common 10x Barcode. Libraries are generated and sequenced and 10x Barcodes are used to associate individual reads back to the individual partitions.

This protocol can be used to generate an enriched T-cell library and/or an enriched B-cell library, and/or a 5’ Gene Expression library from amplified cDNA from the same cells.

Download the 10x Genomics user guide here: VDJ immune profiling and GEX library prep

Uses Chromium Next GEM Single Cell 5′ Library and Gel Bead Kit v1.1 (10x Genomics, 1000165 or 1000167) and
Chromium Next GEM Chip G Single Cell Kit (10x Genomics, 1000120/1000127)
in conjunction with these kits as appropriate:
Chromium Single Cell 5′ Library Construction Kit (10x Genomics, 1000020)
Chromium Single Cell V(D)J Enrichment Kit, Human T Cell (10x Genomics, 1000005)
Chromium Single Cell V(D)J Enrichment Kit, Human B Cell (10x Genomics, 1000016)
Chromium Single Cell V(D)J Enrichment Kit, Mouse T Cell (10x Genomics, 1000071)
Chromium Single Cell V(D)J Enrichment Kit, Mouse B Cell (10x Genomics, 1000072)

10x Genomics cell surface feature/protein library prep

The Chromium Single Cell Feature Barcode technology offers a comprehensive, scalable approach to detect cell surface proteins along with the gene expression and immune repertoire information from the same single cell. This is accomplished by labeling cell surface proteins with antibodies conjugated to a Feature Barcode oligonucleotide, followed by direct capture of the Feature Barcode by the Gel Bead primer. A pool of ~750,000 barcodes are sampled separately to index each cell’s transcriptome and cell surface protein. It does so by partitioning thousands of cells into nanoliter-scale Gel Beads-in-emulsion (GEMs), where all generated cDNA share a common 10x Barcode. Libraries are generated and sequenced and 10x Barcodes are used to associate individual reads back to the individual partitions.

Use this technique to measure both gene and cell surface protein expression in the same cell to identify protein isoforms, detect protein for low abundance transcripts, and further increase phenotypic specificity. This technology can also determine antigen specificity of single T cells with Feature Barcode peptide-MHC multimers to study the dynamic interactions between lymphocytes and antigens. It can also be used to multiplex more than one sample into a single library prep.

This service can be performed in conjunction with the 5′ Next GEM library prep or the VDJ immune profiling (B cell or T cell) library prep, but is not a stand alone service.

Download the 10x Genomics user guide here: Feature Barcode for Cell Surface Protein

Uses Chromium Single Cell 5′ Feature Barcode Library Kit (10x Genomics, 1000080)

10x Genomics ATAC-seq library prep

The Chromium Single Cell ATAC Solution provides a comprehensive, scalable approach to determine the regulatory landscape of chromatin in hundreds to thousands of cells in a single sample. This is achieved by transposing nuclei in a bulk solution; then, using a microfluidic chip and the Chromium Controller, the nuclei are partitioned into nanoliter-scale Gel Beads-in-emulsion (GEMs). A pool of ~750,000 10x Barcodes is sampled to separately and uniquely index the transposed DNA of each individual nucleus. Libraries are generated and sequenced, and 10x Barcodes are used to associate individual reads back to the individual partitions, and thereby, to each individual nucleus.

Download the 10x Genomics user guide here: ATAC-seq user guide

Uses Chromium Next GEM Single Cell ATAC Library & Gel Bead Kit v1.1 (10x Genomics, 1000175/1000176) and
Chromium Next GEM Chip H Single Cell Kit (10x Genomics, 1000161/1000162)