Proprietary Bead Bead Deposition Deposition Load DNA & Enzyme beads into PicoTiter ™ Plate
Dec 20, 2015
Proprietary
Pyrophosphate Sequencing ChemistryPyrophosphate Sequencing Chemistry
Sequencing by synthesis – Cyclic additions of 4 nucleotides: dTTP → dATP → dCTP → dGTP → dTTP …
DNA Polymerase
dNTP
DNA
N Pyrophosphate(PPi)
DNA+(1)
Adenosine Phosphosulfate (APS)
ATP SulfurylasePPi + SO4
2-ATP(2)
dNTP (or ATP)Apyrase
dNMP (or AMP) + 2Pi(4)
N’
ATP CO2 + Oxyluciferin + AMP + PPi +Firefly LuciferaseD-luciferin + O2
(3) Light
Proprietary
Simultaneous Enzymatic Reactions in Simultaneous Enzymatic Reactions in Hundreds of Thousands of Picoliter-size WellsHundreds of Thousands of Picoliter-size Wells
Proprietary
Physico-Chemical Processes on PTP Physico-Chemical Processes on PTP A. Nuc Flow
Nuc Diffusion
Convective transport of nucleotides, APS & D-lucInto flow chamber; diffusive transport into wells
B. Nuc Flow
PPi/ATPCyclic
Reactions
Nuc incorporation on DNA → PPi production → Diffuses to Enzyme beads → Cyclic ATP & PPi generation reactions start → light generation starts
C. Apyrase Flow
Reaction decay
Apyrasediffusion
PPi, ATPdiffusion
PPi, ATP start diffusing out of well; Apyrase flows into chamber & diffuses into well → Neutralizes unincorporated nucs & ATP → Cyclic reaction starts to decay → signal starts to decay
D. Wash Buffer
Reaction ends
Apyrasediffusion
PPi, ATPdiffusion
Wash Buffer flows into chamber; Unused apyrase diffuses out of well → PPi, ATP continue to diffuse out of well → Reaction comes to an end → signal decays to baseline