If you’re about to embark on a high throughput sequencing project, choosing the right sequencing instrument to use is an important consideration. Perhaps you’re replicating a published study or repeating an experiment from previous work and the instrument you plan to use is known. If not, the right sequencing instrument should be based on the sequencing goal you are trying to achieve. Instrument features to take into consideration include: number of reads per run, read length, read type (paired or single end), error type, turnaround time and price. Using Genohub’s Shop by Project page, you can enter the number of required reads or coverage you need and instantly compare instruments, filtering by read length and sorting by turnaround time and price. To get a better idea for the differences between NGS instruments, we’ve generated the following comparison: Table 1.
Certain instruments are ideally suited to specific applications. Illumina instruments are versatile and ideal for a variety of sequencing applications, including: de novo assembly, resequencing, transcriptome, SNP detection and metagenomic studies. The HiSeq and GAIIx instruments are both suited for analyzing large animal or plant genomes. High level multiplexing of samples are possible when analyzing species with a smaller genome size. While the Illumina MiSeq outputs significantly fewer reads (Table 1), its read lengths are significantly longer making it ideal for small genomes, sequencing long variable domains or targeted regions within a genome. The only real limitation to the Illumina platform is its relatively short reads compared to other platforms (Roche 454 and PacBio).
The Ion PGM (Ion Torrent), is ideal for amplicons, small genomes or targeting of small regions within a genome. Its low throughput makes it ideal for smaller sized studies. The Ion Proton however is capable of generating significantly larger outputs (Table 1) making sequencing of transcriptome, exome and medium sized genomes possible.
The PacBio RS/RS II breaks the mold of other short reads high throughput sequencing instruments by focusing on length. The reads, averaging ~4.6 kb are significantly longer than other sequencing platforms making it ideal for sequencing small genomes such as bacteria or viruses. Other advantages include its ability to sequence regions of high G/C content and determine the status of modified bases (methylation, hydroxymethylation) without necessitating the need for chemical conversion during library preparation. The instrument’s low output of reads prevent it from being useful for assembly of medium to large genomes.
The Roche 454 FLX+ is typically used in studies where read length is critical. These include de novo assemblies of microbial genomes, BACs and plastids. It’s long read length has made it a favorite of those examining 16S variable regions and other targeted amplicon sequences. The lower output of the FLX and FLX+ instruments make it less cost-effective for transcriptome or larger genome studies. Roche has announced that it will stop producing the 454 in 2015 and end servicing in mid-2016.
The SOLiD series of instruments are high throughput, generating a large number of short reads. De novo sequencing, differential transcript expression and resequencing are all viable applicaions of the SOLiD platform. The weakness of the platform is its short reads making assembly very difficult.
If you’re still not sure about what NGS instrument to choose for your work, feel free to contact us for our complementary sequencing project consultation.