Genomics FAQs

We will always try and accommodate new protocols that we have not previously done. It makes our work exciting and may benefit other groups eventually. Contact us at ngs-core@jcbc.cam.ac.uk

Please contact us to arrange a handover, and please fill in the online submission form here.

For smaller projects, we can provide a price calculator. We will prepare quotations for larger projects and grant applications. Please contact us at ngs-core@jcbc.cam.ac.uk for details

It will depend on the type of project, but in general we are aiming to have all incoming projects fulfilled within 2-3 weeks. This might be longer at especially busy times.

The NGS facility prioritises projects from the Cambridge Stem Cell Institute, however, we are also accepting a wide range of projects from across the University, and from external contacts.

Once your sequencing run has finished, we will send you information on how to download the data over FTP. The output will be the FASTQ files, and some quality reports and demultiplexing statistics.

Unfortunately, we cannot provide bioinformatic support.

It depends on your individual requirements!
Here are some general guidelines for NGS libraries: https://emea.illumina.com/science/education/sequencing-coverage.html

For bulk transcriptome libraries, we generally sequence 20-50 million reads per sample.

A minimum of 20,000 read-pairs per cell are recommended for libraries constructed with the 10X Genomics single cell kits.

Visium HD samples will be sequenced according to the %coverage of the capture area.

The Novaseq has different sized flowcells which adjust to different output needs. 1.5B flowcells have 2 lanes. 10B and 25B flowcells have 8 lanes.

Here is a table to illustrate the amount of paired end reads (in million) per lane.

• You can book the Bioanalyzers and the tapestation on PPMS. Please ask for training/access if necessary.
• We can offer training or access to the Covaris E220 (https://www.covaris.com/e220 )

We would normally ask you to either replace those samples that failed the QC, or continue the project without them. In some cases, it might be necessary to work with degraded RNA samples, and in those cases we have the appropriate kits available that deal with degraded RNA.

The 10X Genomics Cell Preparation Guide describes best practices and methods for washing, concentrating, resuspending, and counting cells.
Some useful information in this Q&A section.

Cells should be washed a minimum of two times and resuspended in 1X PBS (calcium and magnesium-free) with 0.04% BSA. Solutions should not contain EDTA and surfactants as they will inhibit further reactions. Alternative buffers can be found here.

It is very important that we receive the correct number of cells in the correct volume. For accurate cell counting, please refer to this guide and this page.

10X recommends using cells with a diameter of up to 30µm. For larger cells, it is recommended to use nuclei instead.

A 40 µm Flowmi Cell Strainer can be used to reduce the prevalence of aggregates within a cell suspension. To avoid incorporating noisy data from dead cells in your experiment, 10X Genomics provides a demonstrated protocol for dead cell removal.

Yes, FACS samples are compatible with the 10x Single Cell workflow. It is a good method to enrich for specific cell types based on cell surface markers, and to get a clean cell suspension as it also removes the dead cells and debris. For some tips about best practises go here.

We can either process FFPE samples form human or mouse tissues, or we can process fresh frozen/fixed frozen tissues from any organism with the 3’HD assay. Guidelines about tissue preparation to either of these processes are found here and here. Tissue sectioning, fixation, staining, and imaging can be performed at the JCBC Histology core facility, which you should contact in advance.