REACTION CHAMBER

• US 20100120101A1
• Filed: 01/08/2008
• Published: 05/13/2010
• Est. Priority Date: 01/08/2007
• Status: Active Grant

• Current Assignee: Toxic Reports LLC
• Original Assignee: U.S. Genomics, Inc (Toxic Reports LLC)
• Inventors: Patil, Vishal, Gilmashin, Rudolf, Mollova, Emilia
• Granted Patent: US 8,999,636 B2
• Time in Patent Office: Days
• US Class Current: 435/91.530
• CPC Class Codes: C12N 15/1003 Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor

Abstract

A chamber in which an agent, like genomic DNA, may be harvested and optionally manipulated rapidly (e.g., on the order of a few hours), without shearing or fragmentation.

Claims

1. A reaction chamber for harvesting and/or manipulating intact nucleic acids having a length of at least 50 kilobases.

2. The reaction chamber of claim 1, wherein the reaction chamber harvests and/or manipulates intact nucleic acids having a length of at least 50 kilobases in 12 hours or less.

3. The reaction chamber of claim 1, wherein the reaction chamber manipulates intact nucleic acids having a length of at least 50 kilobases and comprises:

a substrate;

an inlet port positioned to direct fluid flow toward the substrate; and

at least one side port.

55. The reaction chamber of claim 1, wherein the reaction chamber harvests and/or manipulates intact nucleic acids having a length of at least 500 kilobases.

4. A method for manipulating a nucleic acid comprising:

harvesting a nucleic acid from a cell and manipulating the nucleic acid in a single chamber in the absence of shearing;

wherein the nucleic acid is at least 50 kilobases in length.

5. The method of claim 4,

wherein the nucleic acid is harvested and manipulated in 6 hours or less.

6. The method of claim 5, wherein manipulating the nucleic acid comprises hybridizing a sequence specific probe to the nucleic acid.

7. The method of claim 5, wherein manipulating the nucleic acid comprises digestion with a restriction endonuclease.

8. The method of claim 5, wherein manipulating the nucleic acid comprises binding an intercalator to the nucleic acid.

9. The method of claim 5, wherein the nucleic acid is at least 100 kilobases in length, at least 150 kilobases in length, at least 250 kilobases in length, at least 500 kilobases in length, at least 750 kilobases in length, at least 1 megabase in length, or at least 5 megabases in length.

10. The method of claim 5, wherein the nucleic acid is harvested and manipulated in 5 hours or less.

11. The method of claim 5, wherein the nucleic acid is harvested and manipulated in 4 hours or less.

12. The method of claim 5, wherein the nucleic acid is harvested and manipulated in 3 hours or less.

13. A method of separating agents from other constituents in fluid, the method comprising:

flowing fluid toward a substrate in a chamber through a fluid introduction port, the substrate configured to prevent the passage of agents that are larger than a threshold size of the substrate;

flowing a first portion of the fluid through the substrate and out of the chamber;

retaining agents and at least some of the other constituents in proximity to the substrate with flow of the first portion of fluid through the substrate;

flowing a second portion of the fluid through one or more fluid exit ports;

moving at least a portion of the other constituents away from the substrate, into the flow of the second portion of fluid, and out of the chamber through the one or more fluid exit ports to separate the agents from the other constituents.

14. The method of claim 13, further comprising:

flowing fluid that contains the agents into the chamber from the fluid introduction port and through the substrate while preventing flow through the one or more fluid exit ports.

15. The method of claim 14, wherein flowing fluid that contains the agents comprises flowing the fluid that contains the agents through a diffuser to distribute the agents about the substrate.

16. The method of claim 13, further comprising:

flowing the second portion of fluid through the one or more fluid exit ports at an increased rate to remove the agents from the substrate and out of the chamber through the one or more fluid exit ports.

17-35. -35. (canceled)

36. A method positioning agents in a fluid flow, the method comprising:

providing a chamber having a substrate and a diffuser;

flowing a first fluid containing agents through a first fluid port, the diffuser and through the substrate in the chamber; and

flowing a second fluid flow through one or more second fluid ports positioned about the substrate in the chamber, the second fluid flow surrounding the first fluid flow to direct the first fluid flow and any agents contained therein toward a central portion of the substrate.

37-38. -38. (canceled)

39. A system for separating agents from non-agents in a chamber, the system comprising:

a chamber comprising;

a fluid introduction port;

a substrate positioned substantially opposite from the fluid introduction port in the chamber, the substrate configured to prevent the passage of agents that are larger than a threshold size of the substrate;

one or more fluid exit ports positioned peripherally about the chamber between the fluid introduction port and the substrate;

a controller configured to control;

a first flow of fluid into the chamber from the fluid introduction port and through the substrate to hold agents and other constituents at the substrate; and

a second flow of fluid into the chamber from the fluid introduction port and through the one or more fluid exit port and a ratio between the first flow and the second flow allows other constituents to move away from the substrate and into the second flow to separate agents and the other constituents.

40-51. -51. (canceled)

52. A method of positioning agents in a fluid flow, the method comprising:

providing a chamber having a substrate and a diffuser;

flowing a first fluid containing agents through a first fluid port positioned substantially centrally in the diffusion section and through the substrate in the chamber; and

flowing a second fluid flow through a plurality second fluid ports positioned circumferentially about the substrate in the chamber, the second fluid flow circumferentially surrounding the first fluid flow to direct the first fluid flow and any agents contained therein toward a central portion of the substrate.

53-54. -54. (canceled)