Single Molecule, Real-Time (SMRT) Sequencing is the core technology behind PacBio sequencing. In this approach, individual molecules of DNA are sequenced separately from each other within so called "Zero-mode waveguides" (ZMWs) in a sequencing chip called SMRTcell. Inside each ZMW, a DNA polymerase that mediates the sequencing event is able to process an average of 200,000 nulceotides per molecule. Because the DNA becomes a single strand circularized molecule during the process of library preparation, the polymerase can perform multiple passes on the same molecule, depending on the size of the DNA. Therefore, by modulating the average size of the DNA material loaded into the SMRTcell, it is possible also to modulate the number of passes and the number of "subreads" generates from each DNA molecule inside each ZMW. Multiple subreads from the same molecule can be colapsed into a single consensus sequence using the "HIFI" appraoch to produce highly accurate Circular Consensus Sequences (CCS).

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