Sequencing

sstDNA library beads are added to the DNA Bead Incubation Mix (containing DNA polymerase) and are layered with Enzyme Beads (containing sulfurylase and luciferase) onto the PicoTiterPlate™ device. The device is centrifuged to deposit the beads into the wells. The layer of Enzyme Beads ensures that the DNA beads remain positioned in the wells during the sequencing reaction. The bead-deposition process maximizes the number of wells that contain a single amplified library bead (avoiding more than one sstDNA library bead per well).

The loaded PicoTiterPlate device is placed into the Genome Sequencer FLX™ Instrument. The fluidics sub-system flows sequencing reagents (containing buffers and nucleotides) across the wells of the plate. Nucleotides are flowed sequentially in a fixed order across the PicoTiterPlate device during a sequencing run. During the nucleotide flow, each of the hundreds of thousands of beads with millions of copies of DNA is sequenced in parallel. If a nucleotide complementary to the template strand is flowed into a well, the polymerase extends the existing DNA strand by adding nucleotide(s). Addition of one (or more) nucleotide(s) results in a reaction that generates a light signal that is recorded by the CCD camera in the Instrument. The signal strength is proportional to the number of nucleotides, for example, homopolymer stretches, incorporated in a single nucleotide flow.