In February 2001, the sequence of the human genome was published in Nature: the results of a revolutionary project that gave rise to the study of genomics as we know it. Lasting over a decade, this project used a ‘hierarchal shotgun sequencing’ Sanger sequencing approach. The use of this approach has since been overtaken by the development of Next Generation Sequencing (NGS) technologies. Today, NGS platforms – thanks to their massively parallel format, their lower starting material requirements and their increased speed – can produce 20,000 times more data per single run than was produced in the Human Genome Project. A human genome can now be sequenced for less than £1000. The low cost, improved accuracy and increased speed of NGS technologies have fundamentally changed the study of genomics.
Today, NGS is a large and rapidly growing market, worth an estimated $5.3 billion. Illumina is the current market leader, accounting for over 80% of the NGS market. Their platforms make use of sequencing by synthesis with fluorescently labelled nucleotides. For sequencing facility Edinburgh Genomics, which provides sequencing services to researchers both locally and globally, the choice of Illumina as the preferred platform was three-fold. Chief Operating Officer, Joel Fearnley, explained: “In general, their proven technology is robust, the equipment is largely reliable and the reagents perform well. Illumina currently provides the most cost-effective method for whole genome sequencing in humans and other organisms, using the HiSeq X platform which Edinburgh Genomics operates. Other Illumina platforms such as the HiSeq 4000 and the MiSeq offer a wide range of throughput and sequencing strategies, which combined with a large portfolio of methods available to prepare libraries and software to process the data, means that most genomics applications are enabled on Illumina sequencers.”
However, Illumina sequencing often requires large, expensive equipment and time-consuming library preparation. Additionally, Illumina can only offer read lengths of up to 300 bp, although it is possible to obtain read data across much longer fragments on Illumina sequencers using technologies such as 10X Genomics Chromium or Illumina TruSeq Synthetic Long-Reads. Because of this, Illumina may not have a monopoly on the NGS market for much longer. The NGS market is becoming ever more competitive, with users requiring longer read length, with high accuracy and reduced cost.
Dr Maria Doitsidou, Chancellor’s Fellow at the University of Edinburgh, currently employs the Illumina sequencing platform to investigate novel genetic pathways involved in neurodegeneration. Using Caenorhabditis elegans as a model organism, her method involves random mutagenesis to reveal phenotypes of interest, followed by NGS to identify the causal mutations. Dr Doitsidou discussed the developments in NGS technologies that she hopes to see in the next few years: “Further reductions in costs are always desirable. Achieving greater read lengths would also be very convenient, if it was accompanied by reduction in error rates, which normally increase with read length. I would especially like to see costs and error rates drop for single-molecule long-read sequencing applications, to allow easy sequencing of repetitive and complex regions or de novo genome sequencing.”
Dr Doitsidou also commented on the accessibility and speed of current NGS technologies: “What would be most practical for our research is if NGS services were as accessible as Sanger sequencing services are nowadays, with comparably fast turnaround times. That would transform the efficiency of our work, as currently turnaround times for external NGS services are in the range of weeks.”
So how is the market changing to meet these commercial demands? We asked Joel Fearnley for an opinion on the latest NGS developments, and which ones he thought may have the biggest impact on commercially available NGS technologies. He listed: “Illumina’s NovaSeq, for its promise of using any method of sequencing, any species and any depth of coverage at reduced costs; plus PacBio and Oxford Nanopore Technologies, for the introduction of new higher throughput instruments producing very long reads”.
The Genia sequencing platform, recently acquired by Roche, is another nanopore-based sequencing platform of note – using similar technology to Oxford Nanopore Technologies – in which changes in electrical current passing through the nanopore represents the presence of different base pairs. Although this technology is still in the development stage and is not yet commercially available, the principle of nanopore-based sequencing technologies offers the possibility of single molecule sequencing, as well as longer reads and real time output.
Naturally, Edinburgh Genomics has plans to stay up to date with the latest technologies and provide users with services to meet their changing demands. “Edinburgh Genomics already has the latest PacBio Sequel and plans to offer a range of long read services in this quarter,” Joel confirmed. “We tested the MinION but it was not production-ready and we were unable to offer the technology as a reliable service. Oxford Nanopore Technologies now markets the GridION X5 which is of interest due to the higher throughput”. Just sixteen years after the publication of the first fully sequenced human genome, and with longer read lengths, lower error rates and reduced costs on the horizon, it is clear that NGS technologies will continue to develop at an incredible rate in an attempt to meet user needs.
This article was written by Bonnie Nicholson.