Methods for high fidelity production of long nucleic acid molecules with error control

• US 8,263,335 B2
• Filed: 04/25/2011
• Issued: 09/11/2012
• Est. Priority Date: 12/10/2002
• Status: Active Grant

• Current Assignee: Massachusetts Institute of Technology
• Original Assignee: Massachusetts Institute of Technology
• Inventors: Carr, Peter A., Chow, Brian Y., Jacobson, Joseph M., Mosley, David W., Emig, Christopher
• Primary Examiner(s): Lu, Frank W
• Application Number:
  US13/093,807
• Publication Number:
  US 20110201057A1
• Time in Patent Office: 505 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 435/91.51, 435/183, 435/283.1, 435/287.1, 435/287.2, 435/6.1, 435/6.11, 435/6.12, 436/94, 536/23.1, 536/24.3, 536/24.33, 536/25.3
• US Class Current: 435/6.1
• 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 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6846 Common amplification features
• CPC Class Codes: C12Q 1/6848 characterised by the means for preventing contamination or increasing the specificity or sensitivity of an amplification reaction
• CPC Class Codes: C12Q 2521/514 Mismatch repair protein
• CPC Class Codes: C12Q 2537/113 Heteroduplex formation
• CPC Class Codes: C12Q 2537/149 Sequential reactions
• CPC Class Codes: C12Q 2565/513 characterised by the pattern of the arrayed 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

Abstract

A method for synthesizing a nucleic acid having a desired sequence and length comprises providing a solid support having an immobilized nucleic acid, performing a nucleic acid addition reaction to elongate the immobilized nucleic acid by adding a nucleotide or an oligonucleotide to the nucleic acid, determining whether the nucleotide or the oligonucleotide is added to the nucleic acid by detecting whether there is an increase in electrophoretic force applied to the solid support when an electric field and a magnetic field gradient are applied to the support, wherein the increase in electrophoretic force applied to the support is caused by adding the nucleotide or the oligonucleotide to the nucleic acid, repeating the addition reaction and determination steps if the nucleotide or the oligonucleotide is not added to the nucleic acid, and continuing until the immobilized nucleic acid has a desired sequence and length.

Claims

1. A method for determining whether a nucleotide or an oligonucleotide is added to a nucleic acid, the method comprising:

a) providing a solid support having an immobilized nucleic acid, wherein the solid support is a bead or particle;

b) performing a nucleic acid addition reaction to elongate the immobilized nucleic acid by adding a nucleotide or an oligonucleotide to the immobilized nucleic acid;

c) after the nucleic acid addition reaction, determining whether the nucleotide or the oligonucleotide is added to the immobilized nucleic acid by detecting whether there is an increase in electrophoretic force applied to the solid support when an electric field and a magnetic field gradient are applied to the solid support, wherein the increase in electrophoretic force applied to the solid support is caused by adding the nucleotide or the oligonucleotide to the immobilized nucleic acid; and

d) repeating steps b) and c) if the nucleotide or the oligonucleotide is not added to the immobilized nucleic acid.

2. The method of claim 1, further comprising a step of synthesizing a new immobilized nucleic acid having a desired sequence and length by repeating steps b) through d).

3. A method for synthesizing a nucleic acid having a desired sequence and length, the method comprising:

a) providing a solid support having an immobilized nucleic acid, wherein the solid support is a bead or particle;

b) performing a nucleic acid addition reaction to elongate the immobilized nucleic acid by adding a nucleotide or an oligonucleotide to the immobilized nucleic acid;

c) after the nucleic acid addition reaction, determining whether the nucleotide or the oligonucleotide is added to the immobilized nucleic acid by detecting whether there is an increase in electrophoretic force applied to the solid support when an electric field and a magnetic field gradient are applied to the solid support, wherein the increase in electrophoretic force applied to the solid support is caused by adding the nucleotide or the oligonucleotide to the immobilized nucleic acid;

d) repeating steps b) and c) if the nucleotide or the oligonucleotide is not added to the immobilized nucleic acid; and

e) synthesizing the nucleic acid having a desired sequence and length by repeating steps b) through d).