16S Loop Long-reads vs Sanger Sequencing for bacterial isolate identification

Updated: May 22, 2020

If your lab, like many other microbiology labs, has a backlog of isolates sitting at -80°C waiting to be analyzed because the funds and/or time for 16S Sanger sequencing is unavailable, you may be able to clear up space with the Loop 16S Long-read Sequencing Service. Loop's Long-read technology leverages the efficiencies of the Illumina next generation sequencing (NGS) platform but at a much higher sequencing accuracy for more reliable species assignments. In addition, unlike conventional Illumina sequencing which can only sequence 150 bp or 300 bp at a time, Loop's technology provides long-read single molecule sequencing, which easily generates sequence across the entire 16S gene.

So, when does it make sense to choose Loop Long-read sequencing instead of Sanger sequencing for bacterial isolate identification?

The answer to when to choose Loop Long-read sequencing instead of Sanger for identifying isolates depends on why you're using Sanger sequencing instead of NGS.

If you care about accuracy

If you are using Sanger sequencing because you're worried about accuracy, then Loop Long-read technology is a robust alternative. With an error rate of 0.0041%, Loop Long-read accuracy is very similar to what you can achieve with Sanger sequencing, with the added bonus of reads that cover the entire 16S gene. By sequencing the entire 16S gene, you can get more accurate and comprehensive operational taxonomic unit (OTU) assignments for confident, species-level identifications.

If you want species-level assignment

As mentioned at the end of the preceding paragraph, Loop Genomics' ability to sequence the entire genome increases your classification ability. For example, when we use Loop Long-read technology to sequence a microbial population, we can assign >99% of the unique 16S molecules at the species or genus level. Note that this is from only the 16S sequence, which is faster and less expensive than whole genome sequencing (although we can do that, too).

If you want to distinguish the sequence of multiple copies of the 16S gene within your isolate

Full-length 16S sequencing of individual molecules can also deliver something that Sanger sequencing cannot—the ability to differentiate between multiple copies of the 16S gene within an individual bacterium and quantify the relative abundance of each copy. This is because Loop Long-read technology delivers contiguous sequence.

If your isolate is not a pure, single species sample

One area where Sanger sequencing runs into trouble is if your sample is not a pure isolate. This is not a problem for Loop Long-read sequencing, which is frequently used for analyzing microbial populations. We can even tell you how much of your isolate is one species versus another.

If you need affordability

Another reason labs use Sanger sequencing is because, at $10 - $20/read, Sanger sequencing feels like an affordable option. And it is definitely an economical choice if you only have a few isolates to sequence. But if you have hundreds of samples to analyze, that cost increases quite a bit when you factor in labor and materials for getting sequence-ready DNA.

With Loop Long-read sequencing, you can send us tubes or 96-well plates of glycerol stock or resuspended colonies and we will not only deliver your sequence, you'll get bioinformatics support that includes OTU assignment. At $16/sample for 96 samples, the ease and affordability becomes better than Sanger sequencing.

If you need affordable and accurate sequencing over more than just the 16S region

Loop Long-read technology can add even more value to your sequencing projects when you need to sequence multiple regions per isolate. We can even do whole bacterial genomes.

Interested in learning more?

If you'd like to clear out your backlog of isolates and you're ready to learn more, or if you have any questions about our technology or services, you can find answers by emailing us at sales@loopgenomics.com.