The Best of Both Worlds: Precision Sequencing™

You might have seen this term on our website: precision sequencing™. But what does it mean?

Precision sequencing is uBiome’s proprietary (and patented) combination of 16S gene and shotgun metagenomics sequencing. We like to think of it as combining the best of both worlds!

 

DNA Sequencing

Sequencing is the term used by scientists to describe the analysis of large biological molecules, most often DNA. In microbial genomics, DNA sequencing is used to determine which microbial genes are present in a particular sample, such as human stool or a soil sample. Such samples can contain dozens to thousands of microbial species.

Most bacteria and other microbes will not grow inside a laboratory. Until two decades ago, it was therefore very hard to find out which microbial species were present in biological samples. The microscopic world of these tiny organisms remained mostly hidden for scientists, and only bacteria that could be easily grown on a petri dish, such as Escherichia coli, could be studied in detail. The rapid development of molecular techniques such as DNA sequencing, however, made it finally possible to study these elusive, microscopic communities.

 

All About 16S

The most popular technique for the analysis of the human microbiome (and that of animals, plants, and environmental samples) has been 16S rRNA (“16S”) gene amplification and sequencing. The 16S gene is a universal gene present in the genome of all bacteria and archaea. Because it consists of both conserved gene sequences (which are identical in most microbes) and variable regions (which are unique for most microbial groups), this gene serves as a marker for the different species of microbes that are present in a sample.

To analyze the microbial composition of a sample, the 16S gene is amplified using polymerase chain reaction (PCR), a method in which millions of copies of one particular gene are generated using a specific enzyme first isolated from a hot spring in Yellowstone Park. The enzyme is called Taq polymerase, and it was voted Molecule of the Year by Science Magazine in 1989.

After amplification of a specific variable region in all the 16S genes in a biological sample, the copies are sequenced, and the data is compared to large online sequence databases.  Because researchers from all over the world have used this technique, there are millions of different 16S sequences uploaded online. If you compare 16S sequences from your sample to those in databases, most of your 16S sequence reads will have a match to a known bacterium or archaeon.

In other words, 16S amplification and sequencing is very useful if you want to learn about which bacteria are present in a biological sample.

 

All About Full Metagenomic Sequencing

16S sequencing might not be enough to answer many other questions a researcher or doctor might have. For example, “To which antibiotics is this bacterium sensitive?” “Which toxins does this bacterium make?”, or “Can this bacterium synthesize vitamin K?” are questions that can often not be answered using only the 16S gene.

To learn more about what the microbes present in a sample can do, researchers have to sequence all the genes in the sample instead of only the 16S gene. The sequencing of all the combined genes present in a biological sample is called shotgun metagenomics. Using this technique, the isolated collective DNA will be broken into small pieces, and all these short fragments will be sequenced and compared to online databases. Metagenomic sequencing detects a very wide range of genes, including genes that are involved in the synthesis of cell components, metabolic enzymes, toxins, or antibiotic resistance.

Unlike 16S sequencing, which is limited to discovering which microbes are present, metagenomics offers a much more detailed look at the different functions of a microbial community. In other words, while 16S sequencing is great at answering “who is there?”, metagenomics can answer “what can they do?”.  uBiome has been doing full metagenomic sequencing for research purposes and with some of our hundreds of academic partners since our founding in 2012.

There are some downsides to metagenomics, though. In order to find all genes from all microbes in a complex microbiome, you would need to do an unfeasibly large amount of sequencing. A stool sample has hundreds of different bacterial species in various amounts, so finding all genes from a rare species is a tedious task. It would be like finding a needle in the microbial haystack, if you will.

Additionally, the online sequence databases, which are very well stocked with 16S reads, contain fewer reads from other microbial genes. While some genes found with metagenomic sequencing can be easily identified, many others may not find a match. Relying on metagenomic sequencing alone, we would learn more about the presence of certain well-characterized genes, such as toxin or fiber-digesting genes, but we may also end up with many unidentified reads from unknown microbes.  

 

Enter Precision Sequencing™

Precision sequencing provides the perfect balance to answer both questions — “Who is there?” and  “What can they do?” — at the same time. Each sample analyzed with precision sequencing is subjected to both techniques. On one hand, 16S gene amplification and sequencing allows us to give you a deep insight on the presence and percentages of the microbes in your sample. On the other hand, metagenomic sequencing will find a wide range of genes involved in different functions in your microbiome, such as the breakdown of dietary components or drugs or the synthesis of toxins and antibiotic-resistance genes.

Using our patented combination of both techniques, precision sequencing will give you answers about the different kinds of  microbes present in your sample, as well as many of their functions and capacities. This combination will give you both the depth and the width that a single approach could not offer you.

Sometimes you don’t have to choose between two good things. You get to have them both!