Method for analyzing a polynucleotide containing a variable sequence and a set or array of oligonucleotides therefor

• US 6,307,039 B1
• Filed: 02/11/2000
• Issued: 10/23/2001
• Est. Priority Date: 04/07/1995
• Status: Expired due to Term

• Current Assignee: Oxford Gene Technology IP Limited (Sysmex Corporation)
• Original Assignee: Oxford Gene Technology Limited (Sysmex Corporation)
• Inventors: Southern, Edwin Mellor, Pritchard, Clare Elizabeth, Case-Green, Stephen Charles
• Primary Examiner(s): Horlick, Kenneth R.
• Application Number:
  US09/502,778
• Time in Patent Office: 620 Days
• Field of Search: 536/24.3
• US Class Current: 536/24.3
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6834 Enzymatic or biochemical coupling of nucleic acids to a solid phase
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6858 Allele-specific amplification
• CPC Class Codes: C12Q 1/6862 Ligase chain reaction [LCR]
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• 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 2521/101 DNA polymerase
• CPC Class Codes: C12Q 2521/501 Ligase
• CPC Class Codes: C12Q 2525/151 repeat or repeated sequences, e.g. VNTR, microsatellite, concatemer
• CPC Class Codes: C12Q 2533/107 Probe or oligonucleotide ligation
• CPC Class Codes: C12Q 2535/125 Allele specific primer extension
• CPC Class Codes: C12Q 2537/125 Sandwich assay format
• CPC Class Codes: C12Q 2561/125 Ligase Detection Reaction [LDR]
• CPC Class Codes: C12Q 2563/179 the label being a nucleic acid
• CPC Class Codes: C12Q 2565/501 being an array of oligonucleotides
• CPC Class Codes: C12Q 2565/518 : characterised by the immobilisation of the nucleic acid sample or target
• CPC Class Codes: C12Q 2565/537 : characterised by the capture oligonucleotide acting as a primer
• CPC Class Codes: C12Q 2600/156 : Polymorphic or mutational markers

Abstract

A method of analyzing a polynucleotide target involves incubating the target with an oligonucleotide probe, generally an array of immobilised oligonucleotide probes, to form a duplex, and using ligase or polymerase to extend one chain of the duplex. A point mutation or variable number tandem repeat section may be analyzed. Arrays of immobilised oligonucleotides are provided for use in the method.

Claims

1. An array of oligonucleotides, for analysing a polynucleotide target containing a variable sequence, in which each component oligonucleotide i) comprises a sequence complementary to the target including an expected variant of the target, and ii) is tethered to a solid support in a chemical orientation which a) permits duplex formation with the target, and b) permits chain extension only when the sequence of the oligonucleotide matches the variable sequence of the target.

5. An array of oligonucleotides as claimed in claim 1, in which different oligonucleotides occupy different locations and each oligonucleotide has a 3′

nucleotide residue through which it is covalently tethered to a support and a 5′

nucleotide residue which is phosphorylated.

2. A set or array of oligonucleotides, for analysing a polynucleotide target containing a variable number tandem repeat sequence, in which each component oligonucleotide i) comprises a sequence complementary to a part of the target immediately adjacent the repeat sequence, ii) comprises a sequence complementary to the repeat sequence of the target and containing a number of repeats expected in the target, and iii) is configured in a way that a) permits duplex formation with the target, and b) permits chain extension only when the number of repeats in the oligonucleotide equals or is less than the number of repeats in the target.

3. A set or array as claimed in claim 2, wherein each component oligonucleotide iii) is configured in a way that b) permits chain extension:

by ligation only when the number of repeats in the oligonucleotide equals the number of repeats in the target;

or by polymerisation only when the number of repeats in the oligonucleotides is less than the number of repeats in the target.

4. A set or array of oligonucleotides as claimed in claim 2, wherein each component oligonucleotide is tethered to a solid support.

6. A set or array of oligonucleotides as claimed in claim 2, in which different oligonucleotides occupy different locations and each oligonucleotide has a 3′

nucleotide residue through which it is covalently tethered to a support and a 5′

nucleotide residue which is phosphorylated.