Technological Advances Make Genome Sequencing More Efficient and Faster
BY DARRELL E. WARD
Cancer is a disease written in code. That code is represented by four letters – A, C, G and T – with each letter corresponding to one of the four building blocks of DNA. The sequence of these four DNA building blocks, called bases, encodes our genetic information.
Changes in that sequence in individual cells can, over time, cause cells to become cancerous. Rather than dying as they should, the damaged cells live on, proliferate and form tumors. Finally, they acquire the ability to migrate to other organs, where they form metastatic tumors.
Understanding the DNA changes that cause cells to become cancerous is critical for developing new and more effective therapies, overcoming treatment resistance and discovering prognostic biomarkers.
DNA sequencing is a vital technology for identifying the gene changes that occur in cancer cells, and Pelotonia funding has enabled The Ohio State University Comprehensive Cancer Center – Arthur G. James Cancer Hospital and Richard J. Solove Research Institute (OSUCCC – James) to keep pace with important advances in sequencing technology.
The human genome is made up of three billion base pairs divided into 23 chromosomes. Each is a ribbon of bases. The longest of these, chromosome 1, is a string of 246 million base pairs; the shortest, the Y chromosome, has 50 million base pairs. The task of gene sequencing is to identify each base in the chain in the order in which it occurs.
The Human Genome Project used gene sequencing to identify all of the 3 billion bases in the normal human genome. The project ended in 2003, lasted 13 years and involved 18 countries. The sequencing alone cost $400 million. In 2012, Pelotonia funds enabled the OSUCCC – James to upgrade to a second-generation gene sequencer called the HiSeq 2000 from an older, slower predecessor, the GenomeAnalyzer IIA. This year, Pelotonia funding is permitting the upgrade of that system to the HiSeq 2500.
“Pelotonia funding has allowed us to keep up with the latest technology and to democratize high-throughput sequencing by lowering the sequencing cost,” says Pearlly Yan, PhD, technical director of the OSUCCC – next-generation sequencing core. “It also provides options for longer runs and fast turnaround time when they are needed.”
For example, the HiSeq 2500 can operate in regular or in fast mode. In regular mode, the technology can sequence five to six human genomes in about 11 days at 30-fold coverage (a measure of sequencing depth to increase confidence in identifying base alterations), much like the HiSeq 2000. In fast mode, the machine can sequence one human genome in 27 hours at that coverage.
“The upgrade gives us the option to run samples fast, such as the sequencing of clinical samples that need timely results,” Yan says. “The upgrade enables the sequencer to read longer DNA fragments, collecting up to 300 base pairs of data right after the upgrade and 500 base pairs of data in the near future. This is really powerful,” Yan says.
Longer reads during sequencing are important for identifying features such as gene insertions and deletions or major chromosomal changes, but longer reads require more reagent and instrument time, so they are more expensive.
Preparing samples for sequencing is called making DNA or RNA “libraries,” and it is a laborious, time-consuming task when done manually. It involves turning DNA or RNA (which is first converted into DNA) collected from a research model or a patient sample into high-quality DNA fragments that can be sequenced.
Pelotonia funding enabled the OSUCCC – James to purchase a robotic device, called a SciClone, to automate the preparation of DNA libraries for larger sequencing projects, such as transcriptome or exome sequencing or whole genomes.
“The robot runs nonstop and can prepare up to 96 samples at once,” Yan says. “It is not fast, but it is precise and reproducible. And it leaves personnel free to do other things.
“The robotic system enables us to generate DNA libraries efficiently and reproducibly, and with the HiSeq upgrade, we can produce highly consistent data faster. It truly speaks highly of the importance of Pelotonia funding to OSUCCC – James research.”