Patent ID: 11891651
Assignee: ELEMENT BIOSCIENCES, INC.
Field: Measurement (Instruments)
Classification: CPC C  G | IPC C  G

Claim 16:
17. A method for pairwise sequencing, comprising:
a) providing a plurality of immobilized single stranded nucleic acid concatemer template molecules, wherein individual concatemer template molecules lack a scissile moiety that can be cleaved to generate an abasic site in the concatemer template molecule, wherein individual concatemer template molecules in the plurality are immobilized to a first surface primer that is immobilized to a support, and wherein the immobilized first surface primer lacks a nucleotide having a scissile moiety;
b) sequencing the plurality of immobilized concatemer template molecules, thereby generating a plurality of extended forward sequencing primer strands, wherein individual immobilized concatemer template molecules have two or more extended forward sequencing primer strands hybridized thereon;
c) retaining the plurality of immobilized concatemer template molecules and replacing the plurality of extended forward sequencing primer strands with a plurality of forward extension strands by conducting a primer extension reaction with a plurality of soluble amplification primers and a plurality of strand-displacing polymerases to generate a plurality of forward extension strands and a plurality of partially displaced forward extension strands that are hybridized to the immobilized concatemer template molecules to form a plurality of immobilized amplicons, and wherein the primer extension reaction generates a plurality of detached forward extension strands; and
d) sequencing the plurality of immobilized partially displaced forward extension strands, thereby generating a first plurality of extended reverse sequencing primer strands, and sequencing the plurality of immobilized detached forward extension strands, thereby generating a second plurality of extended reverse sequencing primer strands, wherein individual immobilized partially displaced forward extension strands have two or more extended reverse sequencing primer strands hybridized thereon, and wherein individual immobilized detached forward extension strands have two or more extended reverse sequencing primer strands hybridized thereon.