Short cycle methods for sequencing polynucleotides

• US 9,012,144 B2
• Filed: 01/18/2011
• Issued: 04/21/2015
• Est. Priority Date: 11/12/2003
• Status: Expired due to Fees

• Current Assignee: Fluidigm Corporation
• Original Assignee: Fluidigm Corporation
• Inventors: Lapidus, Stanley, Buzby, Philip Richard, Harris, Timothy
• Primary Examiner(s): Whisenant, Ethan C
• Application Number:
  US13/008,182
• Publication Number:
  US 20110152114A1
• Time in Patent Office: 1,554 Days
• Field of Search: 435/6, 435/6.1, 536/23.1, 536/24.3
• US Class Current: 435/6.1
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2527/113 Time

Abstract

The invention provides methods for sequencing a polynucleotide comprising stopping an extension cycle in a sequence by synthesis reaction before the reaction has run to near or full completion.

Claims

1. A method for obtaining sequence information from a target nucleic acid, comprising:

a) immobilizing a plurality of different target nucleic acid/primer duplexes at discrete locations on a surface, wherein target nucleic acid portions of said duplexes each comprise the same adaptor sequence, and wherein said primers are hybridized to said adaptor sequence;

b) contacting said plurality of duplexes with a solution comprising a detectably labeled nucleotide that is not a chain terminating nucleotide and an enzyme under conditions wherein the enzyme catalyzes template-dependent addition to a terminus of at least one of said primers, resulting in the addition of a labeled nucleotide to said primer, wherein the label interferes with progression of the enzyme; and

c) detecting the addition to the terminus of said at least one primer,

wherein said addition is indicative of the presence of a complementary sequence in the target nucleic acid.

2. The method of claim 1, wherein said enzyme is a polymerase.

3. The method of claim 1, further comprising repeating steps b) and c) one or more times.