Single Cell (10X Genomics)

Technology Overview

10X Genomics technology encapsulates single cells into Gel Beads-in-emulsion (GEMs).

The generated cDNA from individual cells will go through library preparation and next-generation sequencing to reveal the gene expression profile on the single cell level.

Our Core currently hosts the Chromium Single Cell Controller instrument. Please feel free to contact us for details of service.

Service Charges and Ordering

Single Cell 3′ Gene Expression Analysis

3′-end gene expression profiling of 500 – 10,000 cells per sample

Single Cell 5′ Gene Expression & Immune Profiling

5′-end gene expression profiling of 500 – 10,000 cells per sample
Assemble, annotate full length T cell and/or B cell V(D)J sequences and call clonotypes
Able to examine gene expression profile and resolve TCR/BCR repertoires from the same sample

Add-on Feature Barcode technology with Single Cell Gene Expression Assays

Enable multiomic profiling by measuring cell surface protein expression or CRISPR perturbations simultaneously from the same cell

Self-service (please enquire)

For non-working hours usage (after 17:30 on weekdays, whole days on weekends)

Service prices

The Core offers full service from cell encapsulation using 10X Chromium Single Cell Controller, subsequent library preparation, Illumina sequencing to data analysis by Cell Ranger. Please contact Dr KWOK, Hin (singlecell10x.cpos@hku.hk / 2831-5483) for project discussion and service quotation.

Prices vary with the following factors:

Number of samples per submission
Applications (e.g. 3′ gene expression, 5′ gene expression, V(D)J analysis, etc.)
Number of target cells (affecting the sequencing cost)
Bioinformatics analysis needs
Affiliated institute (HKU, local academics, overseas academics, etc.)

Service ordering

Please refer to Sample Preparation and Submission section for sample requirements.

Please submit service request through iLab.

Technical Details

Single Cell Encapsulation and Library Preparation

Gene Expression Assays are able to capture 500 to 10,000 cells per sample from a single cell suspension.

Cells will be encapsulated into Gel Beads-in-emulsion (GEMs) by the 10X Chromium Single Cell Controller, followed by reverse transcription and library preparation to become a pool of cDNA libraries.
Transcripts from the same cell will be identified by a 10X barcode, while each of these transcripts will be discernible from one another by a Unique Molecular Identifier (UMI).

Source: 10X Genomics

Sequencing Run

Libraries generated are compatible with Illumina sequencers. In order to read the transcript sequences on one end, and the barcode and UMI on the other end, paired-end sequencing reads are required.
Vendor’s specification recommends getting at least 20,000 paired-end raw reads per cell for gene expression studies*.

* Reads per cell required is very cell-type dependent, it is recommended to obtain more reads per cells at the optimization stage.

Workflow

Sample Preparation and Submission

Cell requirement for 3′ or 5′ gene expression studies

For each sample, users are asked to submit a cell suspension of
– concentration 500 – 1000 cells per μl
– size 30 μm or smaller
– well-dissociated without clumps or doublets
– good viability (>85% viable)
– resuspended in 1x PBS + 0.04% BSA (or other buffers recommended by vendor)

Please refer to vendor’s protocol on sample preparation.

Sample Submission

Please contact our colleagues in advance (via phone/email shown here) before bringing the samples to the Core.

After sample submission date and details confirmed, please submit service request through iLab.

Please submit Single Cell (10X Genomics) sample submission form as attachment in iLab.