Method for a continuous rapid thermal cycle system

• US 8,163,489 B2
• Filed: 04/06/2009
• Issued: 04/24/2012
• Est. Priority Date: 10/06/2006
• Status: Active Grant

• Current Assignee: Applied DNA Sciences Incorporated
• Original Assignee: Vandalia Research, Inc. (Applied DNA Sciences Incorporated)
• Inventors: Murray, Elizabeth, Gregg, Derek Allen, Norton, Michael Louis, Swick, Justin Thomas, Towler, William Ian
• Primary Examiner(s): Kim, Young J
• Application Number:
  US12/384,554
• Publication Number:
  US 20100267091A1
• Time in Patent Office: 1,114 Days
• Field of Search: None
• US Class Current: 435/6.12
• CPC Class Codes: B01L 2300/0832 cylindrical, tube shaped
• CPC Class Codes: B01L 2300/123 Flexible; Elastomeric
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• CPC Class Codes: B01L 7/525 with physical movement of samples between temperature zones
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]

Abstract

Disclosed herein is an efficient, high speed production scale synthesis method for high molecular weight organic substances, such as DNA. The invention includes a method of conducting a polymerase chain reaction which comprises transporting a liquid through polymeric tubing disposed through a first reaction cycle region and at least a second reaction cycle region, each of which regions comprises at least a first and a second temperature zone, the temperature in each zone of said at least second region being substantially identical to the corresponding first and second zones in said first region, wherein said liquid is an aqueous solution comprising polymerase chain reaction reactants and a surface absorbing polymer.

Claims

1. A method of conducting a polymerase chain reaction which comprises transporting continuously a liquid through polymeric tubing disposed through a first reaction cycle region and at least a second reaction cycle region, each of which regions comprises at least a first temperature zone and a second temperature zone, the temperature in each zone of said at least second reaction cycle region being substantially identical to the corresponding first temperature zone and second temperature zone in said first reaction cycle region, wherein said liquid is an aqueous solution comprising polymerase chain reaction (PCR) reactants and a block copolymer of ethylene oxide and propylene oxide dissolved in said aqueous solution.

2. The method of claim 1, wherein said method comprises transporting said liquid through from about 10 to 40 reaction cycle regions, each of which reaction cycle regions comprises at least a first temperature zone and a second temperature zone, the temperature in each zone of said reaction cycle regions being substantially identical to the corresponding zone in said first reaction cycle region.

3. The method of claim 1, wherein said polymeric tubing is flexible polytetrafluoroethylene tubing.

4. The method of claim 1, wherein said liquid is transported into, through and out from said polymeric tubing without physical barriers therein.

5. The method of claim 1, wherein said liquid is a homogeneous aqueous solution.

6. The method of claim 1, wherein each of said reaction cycle regions comprises a first temperature zone, a second temperature zone and a third temperature zone.

7. The method of claim 6, wherein the temperature in said first temperature zone is about 94-96°

C., the temperature in said second temperature zone is about 55-60°

C. and the temperature in said third temperature zone is about 70-73°

C.

8. The method of claim 7 wherein the temperature in said first temperature zone is about 95°

C., the temperature in said second temperature zone is about 57°

C. and the temperature in said third temperature zone is about 72°

C.

9. The method of claim 1 wherein said PCR reactants include Taq DNA polymerase and the concentration of said block copolymer is sufficient to protect said Taq DNA polymerase from deactivation.

10. The method of claim 1 wherein said PCR reactants include Taq DNA polymerase and the concentration of said block copolymer is sufficient to protect said Taq DNA polymerase from deactivation.

11. The method of claim 1 wherein the concentration of said block copolymer is about 1.5 mg/ml to about 100 mg/ml.

12. A method of conducting a polymerase chain reaction which comprises the steps of:

a. transporting an aqueous solution comprising a block copolymer of ethylene oxide and propylene oxide through polymeric tubing followed by;

b. transporting continuously a second aqueous solution comprising PCR reactants through said tubing at temperatures sufficient to induce said polymerase chain reaction.

13. A method of conducting a polymerase chain reaction, which comprises transporting continuously a liquid through polymeric tubing disposed through from about 10 to about 40 reaction cycle regions, each of which reaction cycle regions comprises a first temperature zone, a second temperature zone and a third temperature zone, the temperature in each zone of a reaction cycle region being substantially identical to the corresponding zone in the other reaction cycle regions, wherein said liquid is a homogeneous aqueous solution comprising PCR reactants and a block copolymer of ethylene oxide and propylene oxide dissolved in said aqueous solution.

14. The method of claim 13 wherein the temperature in said first temperature zone is about 94-96°

C., the temperature in said second temperature zone is about 55-60°

C. and the temperature in said third temperature zone is about 70-73°

C. and the concentration of said block copolymer is about 1.5 mg/ml to about 100 mg/ml.

15. The method of claim 14 wherein the temperature in said first temperature zone is about 95°

C., the temperature in said second temperature zone is about 57°

C. and the temperature in said third temperature zone is about 72°

C.

16. The method of claim 14 wherein said liquid is transported into, through and out from said polymeric tubing without physical barriers therein.

17. The method of claim 16 wherein said polymeric tubing is flexible polytetrafluoroethylene tubing.