Pacbio and third generation sequencing

2010s: Third generation sequencing

Since Human Genome Project, the cost of sequencing genomes has decreased more than a thousand-fold.

Since the Human Genome Project the development of newer and better DNA sequencing technologies has led to the cost of sequencing genomes decreasing more than a thousand-fold. No sooner had next-generation sequencing reached the market than a third generation of sequencing was being developed.

SMRT enables scientists to effectively ‘eavesdrop’ on DNA polymerase.

One of these new technologies was developed by Pacific Biosciences (PacBio) and is called Single-Molecule Sequencing in Real Time (SMRT). This system involves a single-stranded molecule of DNA which attaches to a DNA polymerase enzyme. The DNA is sequenced as the DNA polymerase adds complementary fluorescently-labelled bases to the DNA strand. As each labelled base is added, the fluorescent colour of the base is recorded before the fluorescent label is cut off. The next base in the DNA chain can then be added and recorded.

SMRT is very efficient which means that fewer expensive chemicals have to be used. It is also incredibly sensitive, enabling scientists to effectively ‘eavesdrop’ on DNA polymerase and observe it making a strand of DNA.

The advantages of SMRT technologies

SMRT can generate very long reads of sequence of 10-30 kilobases long.

SMRT can generate very long reads of sequence (10-30 kilobases long) from single molecules of DNA, very quickly. Producing long reads is very important because it is easier to assemble genomes from longer fragments of DNA. It also means that complete genome sequences can be obtained without the need for the expensive and time-consuming gap closing methods that other technologies require. With the introduction of such sensitive and relatively cheap sequencing methods scientists can re-sequence genomes that have already been sequenced to achieve a higher level of accuracy. For example, using SMRT, Escherichia coli has now been sequenced to an accuracy of 99.9999 per cent!

With third generation sequencing scientists can now begin to re-sequence genomes to achieve a higher level of accuracy.

SMRT technology also makes it much easier to sequence species that have never been sequenced before – known as de novo sequencing.

A mere decade on from the Human Genome Project – and DNA is now being sequenced far more quickly and efficiently than the early pioneers could possibly have imagined.