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Sequential sequencing

  • US 9,822,408 B2
  • Filed: 03/14/2014
  • Issued: 11/21/2017
  • Est. Priority Date: 03/15/2013
  • Status: Active Grant
First Claim
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1. A method, comprising:

  • a) coupling at least one nucleic acid template to a sequencing platform;

    b) subjecting the at least one nucleic acid template to a first round of sequencing using a first sequencing primer, wherein the first sequencing primer comprises a 3′

    terminal nucleotide which anneals to a first binding site in the at least one nucleic acid template, wherein the first round of sequencing produces a first sequencing read of the at least one nucleic acid template;

    c) removing the first annealed sequencing primer and the first sequencing read from the at least one nucleic acid template; and

    d) subjecting the at least one nucleic acid template to a second round of sequencing using a second sequencing primer, wherein the second sequencing primer comprises a 3′

    terminal nucleotide which anneals to a second binding site in the at least one nucleic acid template which is more than 5 nt away from the first binding site, wherein the second round of sequencing produces a second sequencing read of the at least one nucleic acid template.

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