COMPOSITIONS AND METHODS FOR DETECTING GROUP A STREPTOCOCCI

• US 20090170104A1
• Filed: 10/16/2008
• Published: 07/02/2009
• Est. Priority Date: 11/09/2004
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: POLLNER, Reinhold B., Darby, Paul M.
• Granted Patent: US 8,202,978 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/689 for bacteria

Abstract

Compositions, methods and kits for detecting Group A streptococci. Particularly described are oligonucleotides that are useful as amplification primers and hybridization probes for detecting very low levels of Group A streptococci nucleic acids.

Claims

1. A kit for amplifying and detecting a Streptococcus pyogenes nucleic acid sequence comprising:

a first primer that comprises a 3′

terminal target-complementary sequence and optionally a first primer upstream sequence that is not complementary to said S. pyogenes nucleic acid sequence that is to be amplified, said 3′

terminal target-complementary sequence of said first primer comprising 20 contiguous bases contained within SEQ ID NO;

2, or the full complement thereof, allowing for the presence of RNA and DNA equivalents and nucleotide analogs;

a second primer that comprises a 3′

terminal target-complementary sequence and optionally a second primer upstream sequence that is not complementary to said S. pyogenes nucleic acid sequence that is to be amplified, said 3′

terminal target-complementary sequence of said second primer comprising 26 contiguous bases contained within SEQ ID NO;

1, or the full complement thereof, allowing for the presence of RNA and DNA equivalents and nucleotide analogs; and

a probe that comprises a target-complementary sequence and optionally one or more base sequences that are not complementary to said S. pyogenes nucleic acid sequence that is to be detected, said target-complementary sequence of said probe comprising 13 contiguous bases contained within SEQ ID NO;

3, or the full complement thereof, allowing for the presence of RNA and DNA equivalents and nucleotide analogs.

2. The kit of claim 1, wherein said first primer and said second primer are each up to 60 bases in length.

3. The kit of claim 1, wherein said 3′

terminal target-complementary sequence of said first primer and said 3′

terminal target-complementary sequence of said second primer are each up to 33 bases in length.

4. The kit of claim 3, wherein said 3′

terminal target-complementary sequence of said second primer is up to 28 bases in length.

5. The kit of claim 1, wherein said first primer comprises said first primer upstream sequence.

6. The kit of claim 5, wherein said first primer upstream sequence comprises a promoter sequence for T7 RNA polymerase.

7. The kit of claim 3, wherein said 3′

terminal target-complementary sequence of said first primer is selected from the group consisting of SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, SEQ ID NO;

12 and SEQ ID NO;

13.

8. The kit of claim 4, wherein said 3′

terminal target-complementary sequence of said second primer is selected from the group consisting of SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, and SEQ ID NO;

7.

9. The kit of claim 1, wherein said probe is up to 30 bases in length.

10. The kit of claim 1, wherein said probe comprises said optional one or more base sequences that are not complementary to said S. pyogenes nucleic acid sequence that is to be detected.

11. The kit of claim 10, wherein said probe further comprises a fluorophore moiety and a quencher moiety.

12. The kit of claim 11, wherein said target-complementary sequence of said probe is selected from the group consisting of SEQ ID NO:

23, SEQ ID NO;

24 and SEQ ID NO;

25.

13. The kit claim 1, wherein said probe further comprises a detectable label.

14. The kit of claim 13, wherein said detectable label of said probe is selected from the group consisting of a chemiluminescent label and a fluorescent label.

15. The kit of claim 14, wherein said target-complementary sequence of said probe is selected from the group consisting of SEQ ID NO:

19, SEQ ID NO;

20 and SEQ ID NO;

21.

16. A kit for amplifying and detecting a Streptococcus pyogenes nucleic acid sequence comprising:

a first primer that comprises a 3′

terminal target-complementary sequence and optionally a first primer upstream sequence that is not complementary to said S. pyogenes nucleic acid sequence that is to be amplified, said 3′

terminal target-complementary sequence of said first primer selected from the group consisting of SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, SEQ ID NO;

12 and SEQ ID NO;

13;

a second primer that comprises a 3′

terminal target-complementary sequence and optionally a second primer upstream sequence that is not complementary to said S. pyogenes nucleic acid sequence that is to be amplified, said 3′

terminal target-complementary sequence of said second primer selected from the group consisting of SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, and SEQ ID NO;

7; and

a probe that comprises a target-complementary sequence and optionally one or more base sequences that are not complementary to said S. pyogenes nucleic acid sequence that is to be detected, said target-complementary sequence of said probe selected from the group consisting of SEQ ID NO;

19, SEQ ID NO;

20, SEQ ID NO;

21;

SEQ ID NO;

23, SEQ ID NO;

24 and SEQ ID NO;

25.

17. The kit of claim 16, wherein said first primer comprises said first primer upstream sequence.

18. The kit of claim 17, wherein said first primer upstream sequence comprises a promoter sequence for T7 RNA polymerase.

19. The kit claim 16, wherein said probe further comprises a detectable label.