How Next-Generation Sequencing Works & How Our Services Save Time & Money
No matter what your equipment needs might be, Excedr’s lease program can procure virtually any equipment type and is able to accommodate countless brand preferences your end-user might have. Get in touch today and see how leasing can save you both time and money.
All equipment brands/models are available
The Benefits of Excedr’s NGS Leasing Program:
- Eliminates the upfront cost of purchasing equipment by spreading its cost over time
- Minimizes equipment downtime with included complete repair coverage and preventive maintenance
- Takes advantage of potentially 100% tax deductible* payments, providing you significant cash-savings
- Expedites the administrative work needed for instrument procurement and logistics
- Conserves working capital, enabling you to reinvest in your core business and operations (staffing, inventory, marketing/sales, etc.)
- Accommodates all manufacturer and model preferences
*Please consult your tax advisor to determine the full tax implications of leasing equipment.
When the Human Genome Project (HGP) began in 1990, the objective was to determine the DNA sequence of the entire euchromatic human genome within 15 years.
Many met the project with skepticism, as the huge costs of the project seemed to outweigh the potential benefits. Ultimately, it took researchers 10 years and $3B to complete the project.
However, it was widely considered a huge success, and ushered in a new era of medicine. The project’s completion also led to critical advances in the type of sequencing technology used to sequence DNA.
Several years later, the introduction of next-generation sequencing (NGS) brought a fundamentally different approach to DNA sequencing. It greatly reduced the cost and time it took to sequence an entire human genome, and, through advancements in the technology, allowed for targeted DNA enrichment methods that can perform even higher genome throughput at a reduced cost per sample.
What once took years and millions of dollars to complete now only takes a couple thousand dollars and a day or two, depending on the the platform and size of genome being sequenced.
Since its introduction, NGS technology has been used to determine the sequence of DNA or RNA (RNA-seq) to study genetic variation associated with diseases or other biological phenomena using DNA fragmentation (amplicon sequencing), library preparation, and sequencing, all at a massive scale. It continues to evolve at an unprecedented speed to this day.
Originally referred to as “massively parallel sequencing”, this high-throughput sequencing method allows scientists to:
- Routinely examine and sequence a single genome a large number of times (whole-genome sequencing)
- Observe individual changes in genes
- Study population variations in genes or biomarkers using whole exome sequencing
- Study the microbiome and metagenomics
- Differentiate cancer genomes from healthy genomes at higher rates of accuracy, sensitivity, and speed (making a significant impact on the field of oncology)
- Study the epigenome, providing comprehensive views of epigenetic modifications of many species and cell types.
- Investigate the possibility of personalized medicine
In addition, the development of RNA-seq is due in large part to the success of NGS. It has increased scientists’ and researchers’ understanding of RNA biology, gene expression, and the transcriptome, thanks to innovations in long-read and direct RNA-seq technologies as well as advanced computational tools for data analysis.
Furthermore, it allows for de novo sequencing and assembly, which is used when sequencing a novel genome without a reference sequence available for alignment.
Next-Generation Sequencer Methods, Profiling, Applications, & More
Regardless of the specific method, next-generation sequencers rely on the generation of representative, unbiased sources of nucleic acid templates from the complex genomes being interrogated. Clonal amplification of DNA templates, or single DNA molecules, are sequenced in a massively parallel fashion in a flow cell.
The sequencing is conducted in either a stepwise iterative process or in a continuous real-time manner. In this way, the instruments allow for the sequencing of up to billions of individual DNA templates in a single reaction.
There are several main suppliers of NGS platforms and second generation sequencing platforms. This includes Illumina, Thermo Fisher Scientific, Oxford Nanopore Technologies, Roche, and Agilent, among several others. They all share the same fundamental sequencing process, but with varying technologies.
Let’s take a look at the various methods and technologies being used today.
into digital information (0, 1) on a semiconductor chip, similar to the one you might find in your digital camera. Some systems essentially act as the world’s smallest solid-state pH meter to determine DNA sequences.
The DNA is fragmented, attached to beads, and deposited in millions of wells across the surface of the chip. The wells are then sequentially flooded with one nucleotide after another.
If a nucleotide is incorporated into the strand of bead-bound DNA, a hydrogen ion is given off, a chemical change is measured by an ion sensor beneath the well, and a base is called.
This process takes place in millions of wells simultaneously, enabling sequencing in only a few hours.
DNA ligase is used to determine the underlying sequence of the target DNA molecule. A fluorescence-labeled di-based probe hybridizes to its complementary sequence adjacent to the primed template.
The dye-labeled probe is then joined to the primer following the addition of DNA ligase. Non-ligated probes are washed away, and the ligated probe is identified using fluorescent imaging.
Each base is effectively probed by two independent ligation reactions using two different primers.
Single-Molecule, Real-Time Technology
SMRT technology, in which DNA polymerase attaches itself to a strand of DNA to be replicated, examines the individual base at the point it is attached, and then determines which of four building blocks, or nucleotides, is required to replicate that individual base.
After determining which nucleotide is required, the polymerase incorporates that nucleotide into the growing strand that is being produced.
After incorporation, the enzyme advances to the next base to be replicated and the process is then repeated.
Sequencing by Synthesis Technology
Sequencing templates are immobilized on a proprietary flow cell surface that is designed to present the DNA in a manner that facilitates access to enzymes, while ensuring high stability of surface-bound template and low non-specific binding of fluorescently labeled nucleotides.
Solid-phase amplification creates up to 1,000 identical copies of each single template molecule in close proximity, generating a cluster, and because this process does not involve the positioning of beads into wells or mechanical spotting, much higher densities are achieved.
SBS technology uses four fluorescence-labeled nucleotides to sequence the tens of millions of clusters on the flow cell surface in parallel, using a proprietary reversible terminator-based method.
This enables the detection of single bases as they are incorporated into growing DNA strands. Since all four reversible terminator-bound dNTPs are present during each sequencing cycle, natural competition minimizes incorporation bias.
The result is base-by-base sequencing that enables highly accurate data for a broad range of applications.
Nanopore technology is an exciting new method currently being developed.
The technology records characteristic changes in electric current as nucleic acids pass through a synthetic or protein nanopore and will theoretically allow sequencing of a complete chromosome in one step, without the need to generate a new DNA strand.
NGS vs. Sanger Sequencing
NGS can be compared to a number of existing sequencing technologies, such as microarrays, quantitative PCR (qPCR), and Sanger sequencing. However, NGS and Sanger sequencing are often compared the most. While both are similar in principle, the main distinction between them is each method’s sequencing volume.
Both techniques use DNA polymerase to add fluorescent nucleotides one by one onto a growing DNA template strand in order to identify incorporated nucleotide with a fluorescent tag. However, Sanger sequencing, considered by some to be the golden standard of DNA sequencing technology, can only sequence a single DNA fragment at a time. On the other hand, NGS can can sequence millions of segments per run.
Today, NGS is often paired with Sanger sequencing. It is often used to verify NGS results because of its high levels of accuracy. Both Sanger and next-generation sequencing methods are used in genetic analysis in many of today’s labs, as the two methods can actually support one another, producing more accurate and reliable results.
Bioinformatics & Clinical Applications
While NGS is considered an established analysis technology for research applications across the life sciences, analysis workflows still require substantial bioinformatics expertise.
Bioinformatics, in the context of genomics and molecular pathology, uses computational, mathematical, and statistical tools to collect, organize, and analyze large and complex genetic sequencing data and related biological data.
Some common challenges in setting up large-scale analysis workflows include the appropriate selection of analytical software tools, the speed-up of the overall procedure using HPC parallelization and acceleration technology, the development of automation strategies, data storage solutions, and the development of methods for full exploitation of the analysis results across multiple experimental conditions.
More recently, NGS has begun to expand into clinical environments, where it assists diagnostics, enabling personalized therapeutic approaches. One of the most common NGS assays for cancer patients is targeted panel sequencing, which typically interrogates hundreds of targeted genes.
Sequencers produce vast amounts of quantitative and complex sequencing data that many clinical laboratories rely on to identify genetic alterations of clinical relevance. Creating a resource-intensive data processing pipeline to analyze all the NGS data generated has become necessary in today’s clinical environments.
The Bioinformatics CRO, in parntership with Excedr, offer end-to-end bioinformatics services for labs of all sizes. If you’re looking to lease a next-generation sequencer and foresee the need for a bioinformatics team, look no further. From experimental design through to final analysis, The Bioinformatics CRO can help.
Next-Generation Sequencing Technologies for Lease
If you’re interested in purchasing a next-generation sequencer but wish to avoid spending an enormous amount of capital upfront, consider leasing NGS technologies instead.
Leasing your cutting-edge lab equipment can not provide access to top-of-the-line instrumentation, it can also help you manage cash flow more easily and save room in the budget for other important areas, such as hiring more talented people or buying expensive yet necessary consumables.
Additionally, if you’re considering integrating cell sorting technologies and want to lease a flow cytometer (and use it in conjunction with your sequencer), we can help.
Reach us at +1 888-927-3802 or fill out our contact form and we can discuss your specific needs in detail.
This off-balance sheet financing structure provides three options at the end of the term. The lessee has the option to return the equipment to the lessor, renew at a discounted rate, or purchase the instrument for the fair market value. Monthly payments are also 100% tax deductible which yields additional monetary savings.
If you recently bought equipment, Excedr can offer you cash for your device and convert your purchase into a long-term rental. This is called a sale-leaseback. If you’ve paid for equipment within the last ninety days, we can help you recoup your investment and allow you to make low monthly payments. This also frees up money in your budget rather than tying it down to a fixed asset.
Next-Generation Sequencer Manufacturers & Models on the Market
MiniSeq, MiSeq, NextSeq, HiSeq, HiSeq X, NovaSeq
ThermoFisher & Life Technologies:
Ion Torrent, Ion PGM, Ion Proton, Ion S5, Ion S5 XL
Oxford Nanopore Technologies:
MinION, PromethION, GridION X5, SmidgION
PacBio RS II, PacBio Sequel System
GeneReader NGS System, GeneRead, QIAseq