Massive parallel method for decoding DNA and RNA

• US 10,577,652 B2
• Filed: 11/26/2018
• Issued: 03/03/2020
• Est. Priority Date: 10/06/2000
• Status: Expired due to Term

• Current Assignee: Trustees Of Columbia University In The City Of New York (Columbia University)
• Original Assignee: Trustees Of Columbia University In The City Of New York (Columbia University)
• Inventors: Ju, Jingyue, Li, Zengmin, Edwards, John Robert, Itagaki, Yasuhiro
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US16/200,571
• Publication Number:
  US 20190135851A1
• Time in Patent Office: 463 Days
• Field of Search: 435 61
• CPC Class Codes: C07B 2200/11 Compounds covalently bound to a solid support
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/14 Pyrrolo-pyrimidine radicals
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6872 involving mass spectrometry
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 2525/117 incorporating modified base
• CPC Class Codes: C12Q 2525/186 incorporating a non-extendable or blocking moiety
• CPC Class Codes: C12Q 2535/101 Sanger sequencing method, i.e. oligonucleotide sequencing using primer elongation and dideoxynucleotides as chain terminators
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2565/501 being an array of oligonucleotides
• CPC Class Codes: C40B 40/00 Libraries per se, e.g. arrays, mixtures

Abstract

This invention provides methods for attaching a nucleic acid to a solid surface and for sequencing nucleic acid by detecting the identity of each nucleotide analogue after the nucleotide analogue is incorporated into a growing strand of DNA in a polymerase reaction. The invention also provides nucleotide analogues which comprise unique labels attached to the nucleotide analogue through a cleavable linker, and a cleavable chemical group to cap the —OH group at the 3′-position of the deoxyribose.

Claims

1. A method for sequencing a DNA template of at least 30 nucleotides which comprises the stepwise detection of the identities of nucleotide analogues of adenine, guanine, cytosine, and thymine incorporated into the end of a growing strand of complementary DNA synthesized stepwise using a DNA polymerase in a DNA polymerase reaction, wherein the nucleotide analogues act as reversible terminators in the DNA polymerase reaction and wherein one such nucleotide analogue is any of the following:

2. A method for simultaneously sequencing a plurality of different nucleic acid templates which comprises simultaneously applying the method of claim 1 to the plurality of different nucleic acid templates.

3. A method for sequencing a DNA template of at least 30 nucleotides which comprises the stepwise detection of the identities of nucleotide analogues of adenine, guanine, cytosine, and thymine incorporated into the end of a growing strand of complementary DNA synthesized stepwise using a DNA polymerase in a DNA polymerase reaction, wherein the nucleotide analogues act as reversible terminators in the DNA polymerase reaction and wherein one such nucleotide analogue is any of the following:

4. A method for simultaneously sequencing a plurality of different nucleic acid templates which comprises simultaneously applying the method of claim 3 to the plurality of different nucleic acid templates.