Bead emulsion nucleic acid amplification

Patent metadata

Field	Value
Publication number	US-10982274-B2
Application number	US-201916249681-A
Country	US
Kind code	B2
Filing date	Jan 16, 2019
Priority date	Jan 29, 2003
Publication date	Apr 20, 2021
Grant date	Apr 20, 2021

Abstract

Disclosed are methods for nucleic acid amplification wherein nucleic acid templates, beads, and amplification reaction solution are emulsified and the nucleic acid templates are amplified to provide clonal copies of the nucleic acid templates attached to the beads. Also disclosed are kits and apparatuses for performing the methods of the invention.

First claim

What is claimed is:

1. A composition comprising a population of nucleic acid template-carrying beads, wherein each bead comprises one or more species of template nucleic acid molecules attached thereto,
wherein each bead comprises more than 10,000 amplified copies of each of the one or more species of template nucleic acid molecules attached thereto,
wherein the population of beads are suspended in a microreactor comprising a water-in-oil emulsion, and wherein a plurality of the microreactors includes only one bead.

2. The composition of claim 1 , wherein the attachment of each of the amplified copies of the species of template nucleic acid molecules to the bead is mediated by chemical groups or oligonucleotides that are bound to the surface of the bead.

3. The composition of claim 2 , wherein the attachment of each of the amplified copies of the species of template nucleic acid molecules is a covalent chemical attachment.

4. The composition of claim 2 , wherein the attachment of each of the amplified copies of the species of template nucleic acid molecules is a non-covalent attachment.

5. The composition of claim 1 , wherein each bead comprises a single species of template nucleic acid molecules.

6. The composition of claim 1 , wherein the beads are made from a material selected from the group consisting of cellulose, cellulose derivatives, acrylic resins, glass, silica gels, polystyrene, gelatin, polyvinyl pyrrolidone, co-polymers of vinyl and acrylamide, polystyrene cross-linked with divinylbenzene, polyacrylamide latex gels, polystyrene, dextran, rubber, silicon, plastics, nitrocellulose, natural sponges, control pore glass, metals, cross-linked dextrans, and agarose gel.

7. The composition of claim 6 , wherein each bead is approximately 2 to 100 μm in diameter.

8. The composition of claim 7 , wherein each bead is approximately 10 to 80 μm in diameter.

9. The composition of claim 8 , wherein each bead is approximately 20 to 40 μm in diameter.

Assignees

• Roche Molecular Systems Inc

Inventors

• Berka Jan
• Chen Yi-Ju
• Leamon John H
• Lefkowitz Steven
• Lohman Kenton L
• Makhijani Vinod B
• Rothberg Jonathan M
• Sarkis Gary J
• Srinivasan Maithreyan
• Weiner Michael P

Classifications

• B01L7/52

  with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples · CPC title

• G01N21/6458

  Fluorescence microscopy (fluorescence microscopes per se G02B21/0076 and G02B21/16) · CPC title

• C12Q1/6844

  Nucleic acid amplification reactions · CPC title

• B01L2300/0819

  Microarrays; Biochips · CPC title

• G01N21/253

  for batch operation, i.e. multisample apparatus (analytical automats G01N35/00) · CPC title