RSII Pacific Biosciences

Single Molecule, Real-Time DNA Sequencing System

PacBio RSIIIn November 2011 the LGTC acquired the PacBio RS. In 2013 this was updated to a PacBio RSII. The PacBio is a single molecule sequencer for the sequencing of long DNA fragments (up to 60kb read lengths, with the average of ~14kb).

Technology

Sequencing on the PacBio RS is based on Single Molecule Real Time (SMRT) technology. An active DNA polymerase is immobilized on the bottom of each of the 150.000 wells (ZMW=zero mode wavelength). Each time a nucleotide is incorporated it is held in place by the polymerase and a fluorescent signal is detected, in real time. The incorporated nucleotides are identified by the four different fluorescent labels corresponding to A, T, G and C. The collection of data consists of the recording of a movie of light pulses (pulse data) which is then translated into bases (base data). More information about the technology can be found on the Pacific Biosciences website.

Output

You will receive .pls.h5, .bas.h5, and FASTA files.

Applications

  • De novo assembly of small genomes with 99.9999% accuracy
  • Sequencing of long DNA fragments (structural variation, haplotype data)
  • Targeted resequencing
  • Base modification detection (such as methylation or DNA damage)

Future Applications

  • Clinical diagnostics (Roche)