IP Library › Granted Patent US 7,588,891
Granted Patent B2
US 7,588,891 · App. 11/183,077 · Granted Sep 15, 2009

Invasive cleavage of nucleic acids

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Quick Facts
Patent No.
US 7,588,891
App. No.
11/183,077
Granted
Sep 15, 2009
Kind
B2
Abstract

The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

Claims (23)

1. A method of detecting a target nucleic acid, comprising:

a) exposing a sample comprising a target nucleic acid to a purified thermostable FEN-1 endonuclease from an archaebacterial species selected from the group consisting of Pyrococcus furiosus and Methanococcus jannaschii to generate a cleavage product, wherein said cleavage product is produced from a probe oligonucleotide hybridized to said target nucleic acid to form a cleavage structure comprising a duplex and a flap; and

b) detecting said cleavage product, wherein said cleavage product is indicative of the presence of said target nucleic acid.

2. The method of claim 1 , wherein step a) comprises: exposing said sample to said purified thermostable FEN-1 endonuclease and a polymerase.

3. The method of claim 2 , wherein said polymerase comprises a template-independent polymerase.

4. The method of claim 2 , wherein said polymerase comprises a template-dependent polymerase.

5. The method of claim 1 , wherein said detecting comprises detection of fluorescence.

6. The method of claim 1 , wherein said target nucleic acid comprises synthetic target nucleic acid.

7. The method of claim 6 , wherein said synthetic target nucleic comprises an amplified nucleic acid.

8. The method of claim 7 , wherein said amplified nucleic acid is produced using a polymerase chain reaction.

9. The method of claim 1 , wherein said exposing is conducted under isothermal conditions.

10. The method of claim 1 , wherein said exposing comprises hybridizing first and second nucleic acid molecules to said target nucleic acid.

11. The method of claim 10 , wherein said first and second nucleic acid molecules are capable of hybridizing to a target nucleic acid having a first region and a second region, said second region downstream of and contiguous to said first region, wherein at least a portion of said first nucleic acid molecule is completely complementary to said first region of said target nucleic acid and wherein said second nucleic acid molecule comprises a 3′ portion and a 5′ portion, wherein said 5′ portion is completely complementary to said second region of said target nucleic acid.

12. The method of claim 10 , further comprising hybridizing a stacker oligonucleotide to said target nucleic acid.

13. The method of claim 1 , wherein said detecting comprises detecting a non-target cleavage product.

14. The method of claim 11 , wherein said 3′ portion of said second nucleic acid molecule consists of a single nucleotide not complementary to said target nucleic acid.

15. The method of claim 11 , wherein said 3′ portion of said second nucleic acid molecule comprises a 3′ terminal nucleotide not complementary to said target nucleic acid.

16. The method of claim 11 , wherein said 3′ portion of said second nucleic acid molecule comprises a 3′ terminal nucleotide complementary to said target nucleic acid.

17. The method of claim 11 , wherein said 3′ portion of said second nucleic acid molecule comprises a single nucleotide.

18. The method of claim 11 , wherein a 3′ terminus of said 3′ portion of said second nucleic acid molecule comprises an aromatic ring structure that is not a base.

19. The method of claim 11 , wherein said first oligonucleotide further comprises a 5′ terminal portion that is not complementary to said target nucleic acid.

20. The method of claim 1 , wherein said FEN-1 endonuclease comprises the sequence SEQ ID NO:71.

21. The method of claim 1 , wherein said FEN-1 endonuclease comprises the sequence SEQ ID NO:70.

Assignments (9)
RELEASE OF SECURITY INTEREST Recorded Apr 28, 2026
From: BANK OF AMERICA, N.A., AS COLLATERAL AGENT
To: HOLOGIC, INC., ON ITS OWN BEHALF AND AS SUCCESSOR-BY-MERGER TO DIRECT RADIOGRAPHY CORP.; CYTYC CORPORATION, ON ITS OWN BEHALF AND AS SUCCESSOR-BY-MERGER TO BIOLUCENT, LLC; CYTYC SURGICAL PRODUCTS, LLC, AS SUCCESSOR-BY-CONVERSION TO CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; GEN-PROBE INCORPORATED, ON ITS OWN BEHALF AND AS SUCCESSOR-BY-MERGER TO THIRD WAVE TECHNOLOGIES, INC.; GEN-PROBE PRODESSE, INC.; SUROS SURGICAL SYSTEMS, INC.
Reel/Frame 075566/0039 →
CORRECTIVE ASSIGNMENT TO CORRECT THE INCORRECT PATENT NO. 8081301 PREVIOUSLY RECORDED AT REEL: 035820 FRAME: 0239. ASSIGNOR(S) HEREBY CONFIRMS THE SECURITY INTEREST RELEASE. Recorded Nov 9, 2017
From: GOLDMAN SACHS BANK USA, AS COLLATERAL AGENT
To: HOLOGIC, INC.; BIOLUCENT, LLC; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; SUROS SURGICAL SYSTEMS, INC.; THIRD WAVE TECHNOLOGIES, INC.; GEN-PROBE INCORPORATED
Reel/Frame 044727/0529 →
CORRECTIVE ASSIGNMENT TO CORRECT THE INCORRECT PATENT NO. 8081301 PREVIOUSLY RECORDED AT REEL: 028810 FRAME: 0745. ASSIGNOR(S) HEREBY CONFIRMS THE SECURITY AGREEMENT. Recorded Nov 9, 2017
From: HOLOGIC, INC.; BIOLUCENT, LLC; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; SUROS SURGICAL SYSTEMS, INC.; THIRD WAVE TECHNOLOGIES, INC.; GEN-PROBE INCORPORATED
To: GOLDMAN SACHS BANK USA
Reel/Frame 044432/0565 →
MERGER Recorded Oct 23, 2015
From: THIRD WAVE AGBIO, INC.; THIRD WAVE TECHNOLOGIES, INC.
To: GEN-PROBE INCORPORATED
Reel/Frame 036934/0796 →
SECURITY AGREEMENT Recorded Aug 7, 2015
From: HOLOGIC, INC.; BIOLUCENT, LLC; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; DIRECT RADIOGRAPHY CORP.; GEN-PROBE INCORPORATED; GEN-PROBE PRODESSE, INC.; SUROS SURGICAL SYSTEMS, INC.; THIRD WAVE TECHNOLOGIES, INC.
To: BANK OF AMERICA, N.A., AS COLLATERAL AGENT
Reel/Frame 036307/0199 →
SECURITY INTEREST RELEASE REEL/FRAME 028810/0745 Recorded Jun 4, 2015
From: GOLDMAN SACHS BANK USA, AS COLLATERAL AGENT
To: HOLOGIC, INC.; BIOLUCENT, LLC; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; SUROS SURGICAL SYSTEMS, INC.; THIRD WAVE TECHNOLOGIES, INC.; GEN-PROBE INCORPORATED
Reel/Frame 035820/0239 →
SECURITY AGREEMENT Recorded Aug 1, 2012
From: HOLOGIC, INC.; BIOLUCENT, LLC; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS, LIMITED PARTNERSHIP; SUROS SURGICAL SYSTEMS, INC.; THIRD WAVE TECHNOLOGIES, INC.; GEN-PROBE INCORPORATED
To: GOLDMAN SACHS BANK USA
Reel/Frame 028810/0745 →
TERMINATION OF PATENT SECURITY AGREEMENTS AND RELEASE OF SECURITY INTERESTS Recorded Aug 26, 2010
From: GOLDMAN SACHS CREDIT PARTNERS, L.P., AS COLLATERAL AGENT
To: HOLOGIC, INC.; R2 TECHNOLOGY, INC.; SUROS SURGICAL SYSTEMS, INC.; BIOLUCENT, LLC; DIRECT RADIOGRAPHY CORP.; CYTYC SURGICAL PRODUCTS II LIMITED PARTNERSHIP; CYTYC SURGICAL PRODUCTS LIMITED PARTNERSHIP; CYTYC CORPORATION; CYTYC SURGICAL PRODUCTS III, INC.; CYTYC PRENATAL PRODUCTS CORP.; THIRD WAVE TECHNOLOGIES, INC.
Reel/Frame 024892/0001 →
SECURITY AGREEMENT Recorded Jul 29, 2008
From: THIRD WAVE TECHNOLOGIES, INC.
To: GOLDMAN SACHS CREDIT PARTNERS L.P., AS COLLATERAL AGENT
Reel/Frame 021301/0780 →