Sequencing technology at the cutting edge

Sequencing applications

Here at OGC we utilise both Illumina and PacBio platforms to offer a comprehensive sequencing capability. Both technologies enable rapid and accurate sequencing for genomics, transcriptomics and epigenomics investigations. We continually improve our processes in order to validate and standardise the most robust library preparation procedures to match a range of popular applications. Typical studies include whole genome, exome, amplicon, mRNA and miRNA sequencing.

Illumina’s widely adopted next-generation ‘sequencing-by-synthesis’ technology can be run on one of several instruments, each tailored to different output scales, including very large data yields, and specifications. In the Illumina technology, libraries of short DNA fragments are prepared from DNA or RNA samples and loaded onto glass flow cells, where they are initially amplified into an array of millions of clones or clusters. The sequence of each fragment is read off, base by base, by measuring the fluorescence of each cluster after the synchronous incorporation of labelled reversible terminators corresponding to the DNA base letters A, C, G and T. After each cycle, the fluorescent molecules are removed and the process is repeated, allowing sequence fragments of up to 300 nucleotides to be read.

Our PacBio platform (Sequel IIe) offers the latest in long-read technology and SMRT-Link analysis, enabling the generation of accurate read lengths of up to 25Kb with minimal data processing. In this technology, long, circular DNA libraries are created from either DNA or RNA samples, and sequenced directly without further amplification. Each SMRT (single molecule real time) sequencing cell has an array of nano-fabricated wells, or zero-mode waveguides (ZMWs). Each of these wells can accommodate a single circular template from the library. Subsequently, phospho-linked nucleotides are incorporated one base at a time by an anchored polymerase, with the localised light emission from each event being recorded. Because the template is circular, this process can continue for multiple passes of the library template, which results in a highly accurate consensus sequence for the entire length of the fragment.

Further details about the libraries supported and the available sequencing platforms can be found in the dropdown sections below.

OGC supports a wide-range of library types, which are listed here. These library preps have been perfected over more than a decade of lab operations and automation is in place to support many.

Genomic DNA – can be either high or low complexity and can also be prepared using a PCR-free method to eliminate any potential amplification errors.

Exome/targeted capture – to focus on specific regions of interest using hybridisation probes either commercially available or custom-made. Targeting or other enrichment approaches can be cheaper or more efficient than whole-genome sequencing.

Amplicon – another way of targeting specific regions is through PCR. Generated amplicons can either be sequenced directly or fragmented prior to processing.

Bacterial DNA – this automated prep has been optimised to provide the most cost effective approach for sequencing of large numbers of low-complexity genomes.

16S Amplicons – another way of studying bacterial populations is through analysis of the 16S region in metagenomes.

Read more about our bacterial sequencing approaches: 16S Amplicons and Bacterial DNA.

ChIP – this is optimized for low input samples in order to generate libraries from your ChIP experiment.

PolyA – libraries are prepared from polyadenylated (mature mRNA) transcripts.

Ribosomal/ globin depletion – libraries are prepared from RNA depleted for highly abundant sequences.

3’mRNA – this prep generates short tags close to the 3′ end of the transcripts so that simple gene counts can be performed.

Ultra Low RNA – we can also generate libraries from sorted single cells or single-cell amounts of RNA.

Read more about our PolyA, ribosomal depletion, 3’mRNA and ultra low approaches.

No isolation – enrichment for poly(A)+ sequences or depletion of ribosomal RNA are not suitable for all species and situations. We can also generate libraries without first performing either of these steps.

Virus RNA – we specialise in methods for whole-genome sequencing of clinically important viruses, including HCV and HIV from low-input, low-copy samples such as plasma. Please contact us to discuss your specific needs.

TCR – we can perform a targeted enrichment to allow you to study only the T cell repertoire of your bulk samples.

Small RNA – the discovery and profiling of microRNAs and other small non-coding RNA from any organism, using specialised library preparation methods.

Read more about our small RNA approach.

In addition to our ultra low library preparation capability, we also have a dedicated single cell platform that supports a number of applications: 10X Genomics Chromium.

Single Cell Gene Expression – The Chromium Single Cell 3’ Solution provides high-throughput, single cell expression measurements that enable discovery of gene expression dynamics and molecular profiling of individual cell types. Cell surface protein information can be added with the use of Totalseq Antibodies.

Single Cell Immune Profiling – The Chromium Single Cell V(D)J Solution enables profiling of full-length paired V(D)J transcripts from thousands of lymphocytes.  This solution allows assembly of full-length V(D)J sequences on a cell-by-cell basis, providing high-resolution insights into the adaptive immune system. Gene expression can be combined with immunophenotyping using the Totalseq Antibodies to better describe immune cell subsets and functional states.

Single Cell ATAC – The Assay for Transposase Accessible Chromatin (ATAC) for epigenomic analysis of thousands of individual nuclei provides insight into cell types and cell states for deeper understanding of gene regulatory mechanisms.

Single Cell Multiome ATAC and Gene Expression – Designed to simultaneously profile gene expression and open chromatin from the same cell allowing characterisation of cell types and states, and uncover gene regulatory programs.

Targeted Gene Expression – A defined set of transcripts from single cells or tissue sections can be profiled with Targeted Gene Expression. Customizable, comprehensive gene panels will allow reduced sequencing costs of your samples.

Available panels include Human Pan-Cancer Panel, Human Immunology Panel, Human Gene Signature Panel and the Human Neuroscience Panel. It is also possible to design custom panels and further information can be found on the 10X site.

Visium Spatial Gene Expression – This technology allows mapping of the whole transcriptome with morphological context of the tissue. Oxford Genomic Centre can support library preparation and sequencing of cDNA samples derived from Visium solution.

Feature Barcode Technology
• Antibodies for cell surface protein analysis can be added for an extra layer of information. This solution is compatible with 3 prime and V(D)J Solutions. Individual antibodies or pre-designed panels are available from Biolegend.
• 10X’s CellPlex solution for cell hashing & multiplexing uses a series of oligonucleotides conjugated to a lipid, allowing for subsequent pooling of multiple samples to be run on single channel. Benefits of hashing include a reduction of doublet rate, minimising of batch effects and a cost reduction of complex experiment
• Peptide-MHC multimers to measure antigen specificity. Assess the specificity of the adaptive immune system at single cell resolution with simultaneous capture of paired T cell receptors and peptide-MHC multimers, or immunoglobulin heavy and light chains and their cognate antigens.

Can’t see the library type you’re looking for? OGC also supports its users by QC’ing and sequencing their premade, pooled libraries, allowing you much greater flexibility to push the boundaries with your experiment.

 

We are always willing to discuss needs and alternative approaches. Please refer to our guidelines for sample submission details or contact us for more details.

 

Sequencing Platforms

Illumina NovaSeq6000

The NovaSeq 6000 offers an accelerated pace to high-profile projects through flexible, low-cost and rapid turnaround time sequencing. Combining Illumina’s reliable sequencing-by-synthesis and enhanced high-density patterned clustering technologies this platform offers multiple flow cell types and run configurations to enable throughput scaling to suite a variety of applications. Generating up to 3 Tb and 10 billion read pairs in less than two days the largest S4 flow cell provides the most high-throughput and cost-effective solution for whole genome, exome and transcriptome sequencing.

New projects will be run on the latest chemistry, Illumina v1.5.

IIllumina NextSeq500

The NextSeq 500 instrument is the most flexible Illumina platform for fast-turnaround sequencing of individual projects. With two flow cell configurations, it can produce up to 130M (Mid-Output) or up to 400M (High-Output) reads / read-pairs of 75 or 150 bases and its longest run takes just over a day. Adding flexibility for urgent full-service projects, NextSeq 500 is easy to use and currently forms the core of the bookable self-service runs option for local users.

Illumina NextSeq500

The NextSeq 500 instrument is the most flexible Illumina platform for fast-turnaround sequencing of individual projects. With two flow cell configurations, it can produce up to 130M (Mid-Output) or up to 400M (High-Output) reads / read-pairs of 75 or 150 bases and its longest run takes just over a day. Adding flexibility for urgent full-service projects, NextSeq 500 is easy to use and currently forms the core of the bookable self-service runs option for local users.


Illumina MiSeq

The MiSeq offers highly flexible, high-performance sequencing for smaller projects, including targeted sequencing applications, microbial / viral sequencing and amplicon sequencing.


Illumina iSeq100

Coming soon.

Oxford Nanopore MinION

The MinION was the first sequencing platform to be released by Oxford Nanopore Technologies.

We have a number of MinION devices available for WHG members to run their libraries on.

imagePBS

Pacific Biosciences Sequel IIe

The Sequel IIe is the latest evolution of long read sequencing platforms from Pacific Biosciences. Based on the Single Molecule, Real Time (SMRT) technology, the Sequel IIe can produce highly accurate sequencing data (HiFi sequencing) and it’s the ideal solution for resolving RNA isoforms as each transcript can be sequenced as a single molecule. Among other applications including whole genome sequencing, SMRT analysis can also be implemented to identify bacterial epigenetic modifications.