Method for chemical amplification based on fluid partitioning in an immiscible liquid

• US RE46,322 E1
• Filed: 04/30/2015
• Issued: 02/28/2017
• Est. Priority Date: 03/14/2003
• Status: Active Grant

• Current Assignee: Lawrence Livermore National Security LLC (Government of the United States of America)
• Original Assignee: Lawrence Livermore National Security LLC (Government of the United States of America)
• Inventors: Elkin, Christopher J., Anderson, Brian L., Colston, Bill W.
• Primary Examiner(s): Johnson, Jerry D
• Application Number:
  US14/701,392
• Time in Patent Office: 670 Days
• US Class Current: 1/1
• CPC Class Codes: C07D 473/34 attached in position 6, e.g. adenine
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions

Abstract

A system for nucleic acid amplification of a sample comprises partitioning the sample into partitioned sections and performing PCR on the partitioned sections of the sample. Another embodiment of the invention provides a system for nucleic acid amplification and detection of a sample comprising partitioning the sample into partitioned sections, performing PCR on the partitioned sections of the sample, and detecting and analyzing the partitioned sections of the sample.

Claims

1. An apparatus for nucleic acid amplification of a sample, comprising:

means for partitioning said sample into partitioned sections, wherein said means for partitioning said sample into partitioned sections comprises an injection orifice, and

means for performing PCR on said partitioned sections of said sample.

2. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said injection orifice is an injection orifice that produces microdroplets.

3. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said injection orifice is an injection orifice that injects said sample and a PCR reagent.

4. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said means for performing PCR on said partitioned sections of said sample comprises a continuous tube for circulating said partitioned sections of said sample through a heater to perform PCR.

5. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said means for performing PCR on said partitioned sections of said sample comprises a continuous tube for circulating said partitioned sections of said sample through a heater and cooler to perform PCR.

6. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said means for performing PCR on said partitioned sections of said sample comprises a pump, a continuous tube, and a heater.

7. The apparatus for nucleic acid amplification of a sample of claim 1 including means for detection and analysis of said partitioned sections of said sample comprising a laser and a detector.

8. The apparatus for nucleic acid amplification of a sample of claim 1 including means for detection and analysis of said partitioned sections of said sample comprising a blue laser and a detector.

9. The apparatus for nucleic acid amplification of a sample of claim 1 wherein said means for partitioning said sample into partitioned sections comprises means for separating said sample into immiscible slugs.

10. A method of nucleic acid amplification of a sample, comprising the steps of:

partitioning said sample into partitioned sections, wherein said step of partitioning said sample into partitioned sections comprises flowing said sample through an injection orifice, and

subjecting said partitioned sections of said sample to PCR.

11. A method of nucleic acid amplification of a sample, comprising the steps of:

partitioning the sample into partitioned sections,

wherein the sample comprises a nucleic acid and components for performing nucleic acid amplification,

wherein the step of partitioning the sample into partitioned sections comprises flowing the sample through an injection orifice into an immiscible carrier fluid,

wherein the partitioned sections comprise a gelling agent or a gel, and

subjecting the partitioned sections of the sample to nucleic acid amplification.

12. The method of claim 11, wherein the injection orifice injects the sample and a PCR reagent.

13. The method of claim 11, wherein the injection orifice connects a sample flow pathway to a channel or tube comprising an immiscible fluid.

14. The method of claim 11, wherein the partitioned sections are separated by the immiscible carrier fluid.

15. The method of claim 11, wherein the nucleic acid amplification comprises amplification of a DNA target.

16. The method of claim 15, wherein the partitioned sections contain, on average, a single template of a DNA target, and wherein the single template is amplified within the partitioned sections.

17. The method of claim 15, wherein the partitioned sections are exposed to multiple heating and cooling cycles.

18. The method of claim 17, further comprising moving the partitioned sections through a microchannel.

19. The method of claim 11, further comprising detecting the products of the nucleic acid amplification.

20. The method of claim 19, wherein the detecting comprises confocal imaging.

21. The method of claim 19, wherein the detecting comprises laser excitation.

22. The method of claim 19, wherein the partitioned sections are probed for fluorescent signal.

23. The method of claim 11, wherein the partitioned sections comprise microdroplets or slugs.