What is metagenomic sequencing? You might have heard this term many times in relation to genetics, but what does it actually mean, and why is it important?
To answer this question, it is helpful to break down the term. In the definition of national human genome research institute, metagenomics is the study of the structure and function of entire nucleotide sequences isolated and analyzed from all the organisms (typically microbes) in a bulk sample. Metagenomics is often used to study a specific community of microorganisms, such as those residing on human skin, in the soil, or in a water sample.
Within metagenomic research, the most common approach is shotgun sequencing. Shotgun metagenomic sequencing allows researchers to comprehensively sample all genes from all organisms present in a given complex sample. For example, in research by M. H. Y. Leung et al., titled “Characterization of the public transit air microbiome and resistome reveals geographical specificity,” the metagenomic sequencing technique plays an important role in analyzing the full range of genetic material across all organisms. Unlike other sequencing methods, such as 16S rRNA gene sequencing, metagenomic sequencing takes care of the diversity of organisms and thus allows a large and diverse result. For example, the variety of organisms, their popularity, and the protein involved are all included in the results.
Although metagenomic sequencing has become a well-established technique in contemporary science, its capabilities were not recognized a few decades ago. Our ability to study the environment and biology advances with the development of this DNA sequencing technique. Thus, it is necessary to first examine the evolution of DNA sequencing methods, including the earliest approach, Sanger sequencing.
Developed in the 1970s, Sanger sequencing was the first widely adopted method for determining the nucleotide sequence of DNA. It allowed scientists to read DNA accurately, but only one fragment at a time. Sanger sequencing works by copying DNA strands until chain-terminating nucleotides stop the process, producing fragments of different lengths that can be analyzed to determine the DNA sequence. Although Sanger sequencing remains very accurate, the cost of its labor and time impedes the efficiency of DNA sequencing. In metagenomic studies, this shortcoming is amplified due to the massive amount of data that needs to be processed. In other words, more efficient sequencing methods that maintain high accuracy are necessary.
Next-generation sequencing enables massively parallel sequencing. In NGS, DNA extracted from a sample is first broken into short fragments and attached to a solid surface. These fragments are then copied and sequenced in parallel, producing enormous amounts of genetic data in a single run. Later, the genetic data can be analyzed using computational tools such as QC and FastQC for quality control, Fastp for processing, and MEGAHIT for sequence assembly.
However, NGS also introduced new challenges. The short length of sequencing reads can make genome assembly difficult, and the massive volume of data generated requires advanced computational tools for analysis. Despite these limitations, NGS laid the technological foundation for modern metagenomics and remains one of the most widely used sequencing approaches today.