I've come across a bit of confusion about the initialism NGS, so think it would be a good idea to clarify this term (and similar terms like 2GS, SBS, and HTS) for this site with a bit of discussion.

What are the most common initialisms used to describe different sequencing technologies?

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    $\begingroup$ I think your answer is good, but also feel these terms are mainly used for marketing; much more informative to name the sequencing company and platform. NGS is the worst, some 'NGS' like SOLID and 454 are basically retired $\endgroup$ Commented Jun 3, 2017 at 22:02
  • $\begingroup$ NGS isn't as bad as SMRT, which, despite the expansion of "single-molecule real-time sequencing", is considered specific to PacBio and frowned upon as a tag for nanopore sequencing (which is also single-molecule and real-time). $\endgroup$
    – gringer
    Commented Jun 4, 2017 at 7:41

3 Answers 3


Here are my attempts at definitions:

Sanger: A method of sequencing that depends on chain-terminatiing dideoxynucleotides. This sequencing uses the differential flow of DNA sequences of different lengths through a gel to determine the original DNA sequence, producing a single sequence per reaction container.

NGS: Next-generation sequencing, also referred to as 2GS (second-generation sequencing). This term is used to describe the first wave of sequencing technologies that followed Sanger sequencing technology. The use of NGS has become more confusing with the advent of long-read sequencing, because it's a common assumption that "next-generation" refers to the most recent technology (which is incorrect in this case).

HTS: High-throughput sequencing. This term describes any type of sequencing technology that produces large amounts of data, usually in the form of millions of different sequences produced from the same sequencing run.

SBS: Sequencing by synthesis. This term describes a method of sequencing that depends on the synthesis of [DNA] bases in order for sequencing to be carried out. This definition can extend into long-read sequencing (e.g. PacBio sequencers depend on synthesis during sequencing), but is more typically associated with only the second-generation sequencing technology.

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    $\begingroup$ For me ngs and hts are the same thing - any massively parallel sequencing (anything after sanger). That is further divided to 2nd generations sequencing (short reads) and 3rd generation sequencing (long reads). Sequencing by synthesis can by whatever sequencing that uses template and synthesis of free nucleotides (454, Illumina, PacBio). $\endgroup$ Commented Jun 3, 2017 at 23:27
  • $\begingroup$ If you have to divide "next-generation" into multiple generations, it's probably not a good definition. $\endgroup$
    – gringer
    Commented Jun 4, 2017 at 0:41
  • $\begingroup$ Some people will distinguish the ABI SOLiD and Nanostring Hyb+Seq sequencing technologies as "sequencing by ligation" rather than "sequencing by synthesis". I currently lump these all together as SBS because I haven't found a better descriptive name for the collective term. $\endgroup$
    – gringer
    Commented Jun 4, 2017 at 0:44
  • $\begingroup$ I did bit of digging about "Sequencing by synthesis" : it is defined on an Illumina flyer, page 15, it is defined though florescent died nucleotides... PacBio write on their web : "Also, while it is a form of ‘sequencing by synthesis’, it operates in real-time rather than incorporating a single base (or type of base) at a time." $\endgroup$ Commented Jun 4, 2017 at 10:36
  • $\begingroup$ It's not quite "real-time" sequencing, as the base-calling is done by decoding fluorescence "movies" for each photodetector (ZMWG)... but then again nothing is truly real-time unless it's detecting and calling single bases at the same time. $\endgroup$
    – gringer
    Commented Jun 4, 2017 at 10:45

I am not sure how appropriate it is at this point to still refer to sequencing as next-generation sequencing. The leading NGS technology is Illumina/Solexa that has been around for over 10 years at this point. 454 was around even earlier. It's not really "next" at this point.

Opinions aside, I would also add "third-generation sequencing" to that list, referring to long read technologies like PacBio and Oxford Nanopore. See this question on Biology SE for more details.

  • $\begingroup$ I have a bit of trouble classifying PacBio in a sequencing generation. It is a model-based sequencing technology (dependent on a polymerase and pre-mixed nucleotides for sequencing), but produces single-molecule reads sequentially over the course of a run. I therefore consider it to be somewhere between second and third generation sequencing technologies. $\endgroup$
    – gringer
    Commented Jun 5, 2017 at 21:01

The terms "second-generation sequencing", "third-generation sequencing", "short-read sequencing", and "long-read sequencing" are used commonly but (as far as I know) lack commonly used acronyms/initialisms.

In the forensic genetics community, the acronym MPS is used commonly to refer to "massively parallel sequencing." This definitely includes 2G/short-read sequencing (such as Illumina). It's less clear whether MPS includes 3G/long-read sequencing such as PacBio or Oxford Nanopore, but since these are rarely used in a forensics context it's a moot point.

Of course, it's important to acknowledge (as @burger has done) that there is a lot of imprecision and opinion regarding the appropriate scope of each term. Some folks seem to want a term to refer to everything post-Sanger, which includes an incredible diversity of platforms with respect to sequencing technology, read length, throughput, and error profiles. Some very reasonable questions should make us think seriously about our chosen terminology: How long do (for example) Illumina reads have to get before they're no longer classified as short reads? How much throughput is required of a sequencer to be classified as "high" throughput? When do these distinctions really matter?


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