10X Chromium Single Cell ATAC Cromatin Profiling
The Chromium Single Cell ATAC Solution allows the profiling of cellular chromatin from individual cell nuclei ranging from 500 to 10,000 nuclei per sample and up to 80,000 nuclei per run. The Single Cell ATAC (Assay for Transposase Accessible Chromatin) Solution accelerates the understanding of the regulatory landscape of the genome providing insights into cell variability. High throughput profiling of chromatin at a single cell level allows researchers to see how chromatin compaction and DNA-binding proteins regulate gene-expression. The 10X Genomics ATAC workflow allows your project to reveal insights into chromatin accessibility at a single cell level.
- Discover Cellular heterogeneity derived from epigenetic variability
- Better understand the gene regulatory networks that are upstream of gene expression
- Define cell types and states for lineage and developemental program tracing
- Study developemental plasticity
- Accelerate biomarker discovery
- Investigate cell lines, primary cells, fresh and cryopreserved samples
Areas of Research
- Stem Cell / Developmental Biology
Single cell ATAC libraries prepared using the 10X Chromium Controller are Illumina compatable and will be sequenced using the NovaSeq 6000. A 100 cycle sequencing profile is used to sequence 10X single cell ATAC libraries. For a typical single cell ATAC project targeting 10,000 nuclei @ 25,000 read pairs per nuclei, a full S1 flow cell will be required to run 6 multiplexed libraries or 3 multiplexed libraries on a full SP flow cell.
The success of a single cell ATAC sequencing project is determined by the quality of the cell nuclei preparation provided to Genomics Core. Researcher must coordinate their sample delivery with core personnel at least 2 weeks prior to submission. The nuclei sample must be submitted as a completely dissassociated nuclei suspension derived from cells with a high viability percentage of >80%. Cryopreserved cells are acceptable for nuclei isolation. For cell samples undergoing Flow Cytometry sorting prior to nuclei isolation, additional coordination between the Flow Cytometry Core and the Genomics Core will be required. The concentration and viability of each nuclei suspension will be assayed using the Contess II FL Automated Cell Counter. An assessment of nuclei viability at a high magnification of ≥ 50X is requested by the Genomics Core to ensure that blebbing of the nuclei is not present or prevalent (magnified images are appreciated).
- Plan your nuclei preparation time frame to conform with the predetermined Genomics Core submission schedule. Contact the Genomics Core Staff 2 weeks prior to your submission date to coordinate your ATAC-seq project.
- Fresh cells, tissue or cryopreserved cells can be used for nuclei isolation.
- Primary/fragile cells may have high amounts of ambient/background DNA. Treating the cells with DNase I prior to nuclei isolation can remove ambient DNA improving the quality of Single Cell ATAC libraries. See 10X Demonstrated Protocol : Nuclei Isolation for Single Cell ATAC Sequencing in the links below.
- Solid tissues and other large cell aggregates must be disassociated using mechanical or enzymatic disassociation.
- It is recommended that a trial nuclei isolation procedure be performed before submitting a nuclei prep to the Genomics Core to determine the nuclei concentration and clarity of the suspension.
- Critical : Resuspend nuclei isolates in 1x Nuclei Buffer using nuclease free water as the diluent prior to sample submission. Nuclei Buffer 20x (10X Genomics PN-2000153 / 2000207). Nuclei Buffer is optimized for the transposition and barcoding steps in the Single Cell ATAC protocol.
- Minimize delay between nuclei preparation and submission. Keep nuclei on ice.
- To ensure a well singulated nuclei suspension free from cell debris, nuclei straining can be employed. Ensure that pore size is larger than the nuclei diameter but small enough to catch clumps and debris. Account for volume loss associated with the straining process and repeat the nuclei count to correct for loss.
- Submit nuclei samples with a concentration of 3,080 - 7,700 nuclei/ul (3 - 7.7 million nuclei/ml) to target 10,000 nuclei for interrogation.
- An aliquot of 1X Nuclei Buffer can be provided by the Genomics Core personnel when coordinating your Single Cell ATAC project with the core.
Supporting documents and video links for cell preparation, nuclei isolation and project planning
- 10X Video: Chromium Single Cell ATAC Solution
- 10X Demonstrated Protocol: Nuclei Isolation for Single Cell ATAC Sequencing
- 10X Demonstrated Protocol: Nuclei Isolation from Mouse Brain Tissue for Single Cell ATAC Sequencing
- 10X Document: Profiling Chromatin Accessibility at a Single Cell Resolution
- 10X Application Note: Single Cell ATAC-seq for Characterization of Complex Bilogical Systems
- 10X Genomics Cell Preparation Guide
- 10X Genomics Tumor Dissociation for Single Cell RNA Sequencing Guide
- 10X Single Cell Community Forum