Plasmid purification system and method

• US 5,139,637 A
• Filed: 03/13/1991
• Issued: 08/18/1992
• Est. Priority Date: 03/13/1991
• Status: Expired due to Term

• Current Assignee: MacConnell, William P.
• Original Assignee: MacConnell, William P.
• Inventors: MacConnell, William P.
• Primary Examiner(s): Niebling, John
• Assistant Examiner(s): STARSIAK, JOHN S
• Application Number:
  US07/668,856
• Time in Patent Office: 524 Days
• Field of Search: 204/182.8, 204/182.1, 204/182.7, 204/182.9, 204/299 R, 204/301
• US Class Current: 204/466
• CPC Class Codes: G01N 27/4473 by electric means
• CPC Class Codes: G01N 27/44756 Apparatus specially adapted therefor

Abstract

An apparatus for the purification of DNA and the like comprises a housing having walls forming a reservoir having a plurality of chambers for containing a buffer solution means for circulating a buffer through the reservoir, a disposable cassette within said housing having first means including a gel for defining a first path extending between a first pair of the chambers, a well for introducing a bacterial sample into the path at one end thereof, and a second path intersecting the first path via an elution window at one end, having a collection window at the other end and extending between a second pair of the chambers, and an electrical circuit for selectively applying an electrical potential along each of the paths for selectively moving a plasmid first along the first path from the bacterial well to the elution window, then along the second path to the collection window at the end thereof.

Claims

1. An apparatus for the purification of DNA and the like comprising:

a housing forming a reservoir having a plurality of chambers for containing a buffer solution;

means for circulating a buffer through said reservoir;

disposable means positionable within said housing and having first means including a gel for defining a first path extending between a first pair of said chambers, means for introducing a bacterial sample into said path at one end thereof, and second means for defining a second path intersecting said first path via an elution window at one end, having a collection window at the other end and extending between a second pair of said chambers; and

means for selectively applying an electrical potential along each of said paths for selectively moving a plasmid first along said first path from said bacterial sample to the intersection of said first path then along said second path to said collection window at the end thereof.

2. A purification apparatus according to claim 1 wherein said gel is an electrophoretic agarose gel.

3. A purification apparatus according to claim 1 wherein said gel is an agarose-acrylamide double phase gel.

4. A purification apparatus according to claim 3 wherein said gel has an approximate one percent agarose phase extending from said inlet means part way to said elution window, and an approximate five percent acrylamide phase extending from said agarose phase to said elution window.

5. A purification apparatus according to claim 1 wherein said disposable means comprises a body formed of a non-conductive material with upper and lower spaced apart panels for containing said gel and defining said first path, wall means extending upward from around a window in said upper panel for defining said inlet means, upstanding walls surrounding said elution window for defining at least part of said second path, and said collection window comprises a membrane covering an opening in said upstanding walls.

6. A purification apparatus according to claim 5 wherein gel is an electrophoretic agarose gel.

7. A purification apparatus according to claim 5 wherein said gel is an agarose-acrylamide double phase gel.

8. A purification apparatus according to claim 7 wherein said gel has an approximate one percent agarose phase extending from said inlet means part way to said elution window, and an approximate five percent acrylamide phase extending from said agarose phase to said elution window.

9. A purification apparatus according to claim 5 wherein said disposable body is formed of a plurality of said first paths and a plurality of said second paths.

10. A purification apparatus according to claim 5 wherein said membrane of said collection window comprises a dialysis membrane extending at an angle of approximately forty-five degrees to the horizon, disposable body is formed of a plurality of said first paths and a plurality of said second paths.

11. An apparatus for the purification of DNA and the like comprising:

a generally box-like housing forming a reservoir having a plurality of chambers for containing a buffer solution;

means for circulating a buffer through said reservoir;

a disposable cassette detachably mountable within said housing and having a pair of vertically spaced horizontally extending walls, including an electrophoretic agarose gel disposed therebetween for defining at least a part of a first path extending generally horizontally between a first pair of said chambers, an inlet well for introducing a bacterial sample into said path at one end thereof, and vertically aligned openings in said walls defining an elution window, a microchamber having a collection window and communicating with said elution window for defining a second path intersecting said first path via said elution window and extending between a second pair of said chambers; and

means for selectively applying an electrical potential along each of said paths for selectively moving a plasmid first along said first path from said inlet well to said elution window, then along said second path to said collection window at the end thereof.

12. A purification apparatus according to claim 11 wherein said disposable cassette comprises a body formed of a non-conductive material wherein said walls comprise upper and lower spaced apart panels for containing said gel and defining said first path, wall means extending upward from around a window in said upper panel for defining said inlet well, upstanding walls surrounding said elution window for defining at least part of said second path, and said collection window comprises a dialysis membrane covering an opening in said upstanding walls.

13. A purification apparatus according to claim 12 wherein said gel is an agarose-acrylamide double phase gel.

14. A purification apparatus according to claim 13 wherein said gel has an approximate one percent agarose phase extending from said inlet means part way to said elution window, and an approximate five percent acrylamide phase extending from said agarose phase to said elution window.

15. A purification apparatus according to claim 12 wherein said disposable cassette includes a plurality of vertically extending walls disposed between said vertically spaced horizontally extending walls forming a plurality of said first paths and a plurality of said second paths.

16. A purification apparatus according to claim 12 wherein said membrane of said collection window comprises a dialysis membrane extending at an angle of approximately forty-five degrees to the horizon.

17. A process for the purification of DNA and the like comprising the steps of:

selecting a housing having walls forming a reservoir having a plurality of chambers for containing a buffer solution;

circulating a buffer through said reservoir;

selecting and positioning disposable means within said housing and having first means including a gel for defining a first path extending between a first pair of said chambers, means for introducing a bacterial sample into said path at one end thereof, and second means for defining a second path intersecting said first path via an elution window at one end, having a collection window at the other end and extending between a second pair of said chambers;

introducing a bacterial sample into said path at one end thereof; and

selectively applying an electrical potential along each of said paths for selectively moving a plasmid first along said first path from said bacterial sample to said elution window, then along said second path to said collection window at the end thereof.

18. A purification process according to claim 17 wherein said gel is an electrophoretic agarose gel.

19. A purification apparatus according to claim 17 wherein said gel is an agarose-acrylamide double phase gel.

20. A purification apparatus according to claim 19 wherein said gel has an approximate one percent agarose phase extending from said inlet means part way to said elution window, and an approximate five percent acrylamide phase extending from said agarose phase to said elution window.