Pacific Biosciences of California, Inc.
Nanopore sequencing methods

Abstract:

Methods are provided for sequencing of nucleic acid templates using nanopores. A polymerase-nucleic acid complex having a strand displacing polymerase is provided. The nucleic acid in the polymerase-nucleic acid complex is a circular nucleic acid template. Components required for nucleic acid synthesis are provided whereby the polymerase produces a nascent strand that passes through the nanopore while the nascent strand is produced. Current through the nanopore over time is measured allowing for the determination of the sequence of the nascent strand as it translates through the pore using the measured current over time. Because the polymerase can proceed around the circular template multiple times, multiple reads corresponding to the same sequence in the circular template can be obtained in one sequencing process. Where modified bases are present in the template, the method can be used to determine the presence and position of the modified bases using the kinetics of the polymerase.