Overview - A multiplatform approach for sequencing
A multiplatform approach in next-generation sequencing (NGS) involves the combined analysis of sequencing reads generated on two or more different sequencing platforms. Most often researchers that require very long read lengths with very low error rates opt for this sequencing strategy. Having physical access to a range of sequencing instruments and the technical knowledge to handle various sequencing technologies are both key elements in properly applying a hybrid sequencing approach.
One successful example of merging sequencing reads from different sequencing systems is the combination of PacBio’s long reads with Illumina’s low-error shorter reads. This cross-platform method is popular for finishing eukaryotic whole genomes, for resolving complex genomic regions, for phasing haplotypes and for characterising transcripts with high sequence identity.
Applications - The advantages of using cross-platform sequencing
A hybrid approach to next-generation sequencing can lead to:
- Improved error correction
- Decreased number of misassemblies
- Higher number of correctly mapped genomic regions
- Complete genome assembly
- Analysis of full-length cDNA sequences
- Genotyping with increased accuracy
- Identification of novel conformation changes between cancer cells
Sequencing technologies and instruments
GATC Biotech houses all state-of-the-art sequencing platforms in its own laboratories. The industry-leading Illumina instruments and the popular PacBio platform can be used alone or in combination to meet the sequencing requirements of a wide variety of projects.
Next-generation sequencing technologies
GATC Biotech was the first multiplatform sequencing service provider in the world by initially offering NGS services on both Roche and Illumina systems.
GATC Biotech started out providing NGS services based on the pyrosequencing technology of Roche’s sequencing systems. Pyrosequencing relies on chemiluminescence detection of pyrophosphates that are released during DNA synthesis. The library DNAs with specific adaptors are denatured into single strands and enriched by amplification beads with emulsion PCR. During subsequent DNA synthesis, whenever the DNA polymerase incorporates a nucleotide that is complementary to the DNA template, a pyrophosphate molecule is released and converted to ATP. The ATP molecule provides energy for an enzymatic reaction that generates light. Because nucleotides are added in a stepwise process and their identities are known, the sequence of the template can be determined.
GATC Biotech then quickly mastered another sequencing-by-synthesis (SBS) technology that was initially developed by Solexa and later by Illumina. The method involves fixing a library with adaptors, denaturing to single strands and bridge amplifying to form clusters of DNA fragments. The library is then spliced into single strands and sequenced. During sequencing, each of the four kinds of nucleotides that have been modified to contain a cleavable fluorescent dye and a removable blocking group are added sequentially. Upon nucleotide incorporation, a fluorescent signal is emitted and recorded in order to determine the DNA template sequence.
Since then, GATC Biotech has also incorporated a third-generation sequencing approach, the single-molecule real-time (SMRT) technology. This sequencing process uses SMRT cells embedded with millions of zero-mode waveguides (ZMWs). A single DNA polymerase is attached to the bottom of a ZMW with a single DNA molecule serving as a template. The structure of the ZMW creates an observation volume that is so small that it only allows the detection of a single nucleotide incorporation event. For the sequencing reaction, four nucleotides are used, each labelled with a different fluorescent dye. When a base is incorporated by the polymerase, the fluorescent tag is cleaved off and the emission signal is recorded in real time. The time difference between two incorporation events can provide additional information about the structural variance and epigenetic modifications of the sequence.
Next-generation sequencing systems
Several high-throughput NGS platforms using corresponding sequencing technologies exist. The most popular massively parallel sequencing machines include Illumina’s MiSeqs and Illumina’s HiSeqs, which apply SBS chemistry. For long read lengths, the PacBio platforms, which are based on SMRT technology, are the systems of choice.
Scientific expertise: Illumina MiSeq, Illumina HiSeq and PacBio RS II
In 2006, GATC Biotech became the first sequencing service provider to offer a multiplatform sequencing approach with the leading technologies at that time. In 2010, GATC Biotech was the first company to install the Illumina HiSeq 2000 for commercial use, followed by the PacBio RS II one year later. With years of experience handling the most popular sequencing platforms, the team is fully prepared to advance projects that require either one or both of its leading sequencing systems.
Find here, a list of selected research articles supported by GATC Biotech’s sequencing products, including publications that have applied a multiplatform sequencing approach.
Products related to multiplatform sequencing
Convinced a cross-platform approach will benefit your next-generation sequencing project? Try our NGSELECT product for maximum flexibility in the generation and analysis of sequencing reads.
Interested in seeing how Illumina’s platforms can advance your research? Rely on one of our INVIEW pre-designed services for exome, as well as for transcriptome, microbiome or epigenome analysis.
Ready to explore with PacBio’s ultra-long reads? See how you can use one of our standardised INVIEW products to enhance your de novo genome finishing, microbiome (INVIEW MICROBIOME HIGH SPECIFICITY) analysis with the SMRT technology. Take advantage of our all-in-one packages offering DNA isolation, library preparation, sequencing and BioIT analysis all performed by a team of multi-disciplinary experts.
Please contact us with any questions on how our sequencing platforms can be used to achieve your project objectives.
Further reading on multiplatform sequencing
Jeong, H., Lee, D.H., Ryu, C.M., Park, S.H. Toward Complete Bacterial Genome Sequencing through the Combined Use of Multiple Next-Generation Sequencing Platforms. J Microbiol Biotechnol. 26(1):207-12 (2015).