Hybridization assay probes and methods for detecting the presence of Neisseria meningitidis subtypes A,C and L in a sample

• US 20030198984A1
• Filed: 02/12/2003
• Published: 10/23/2003
• Est. Priority Date: 06/07/1995
• Status: Abandoned Application

• Current Assignee: Gary Bee, Sherrol Mcdonough, Yeasing Yang
• Original Assignee: Gary Bee, Sherrol Mcdonough, Yeasing Yang
• Inventors: Yang, Yeasing, McDonough, Sherrol, Bee, Gary
• Application Number:
  US10/365,034
• Publication Number:
  US 20030198984A1
• Time in Patent Office: Days
• US Class Current: 435/6.15
• CPC Class Codes: C12Q 1/689 for bacteria

Abstract

The present invention discloses hybridization assay probes, amplification primers, nucleic acid compositions and methods useful for detecting Neisseria nucleic acids. Hybridization assay probes and amplification primers that selectively detect Neisseria meningitidis and distinguish those Neisseria meningitidis from Neisseria gonorrohoeae are disclosed. Other hybridization probes selectively detect Neisseria gonorrohoeae and not Neisseria meningitidis are also described.

Claims

1. A hybridization assay probe for use in detecting the presence of Neisseria meningitidis in a sample, said probe comprising a base sequence region up to 100 bases in length which is able to form a detectable hybrid with a first target nucleic acid region present in nucleic acid from or derived from Neisseria meningitidis subtypes A, C and L and which is unable to form a detectable hybrid with nucleic acid from Neisseria gonorrhoeae under stringent hybridization assay conditions, wherein said first target region has a sequence selected from the group consisting of SEQ ID NO. 11, SEQ ID NO. 15, SEQ ID NO. 25 and SEQ ID NO. 27.

2. The probe of claim 1, wherein the base sequence of said base sequence region is at least 80% complementary to the base sequence of said first target region.

3. The probe of claim 1, wherein the base sequence of said base sequence region is fully complementary to the base sequence of said first target region.

4. The probe of claim 1, wherein the base sequence of said probe is at least 80% complementary to the base sequence of said first target region.

5. The probe of claim 1 further comprising a detectable label.

6. A composition comprising a nucleic acid hybrid formed between said probe and said first target region of claim 1.

7. A composition comprising a nucleic acid hybrid formed between said probe and said first target region of claim 2.

8. A composition comprising a nucleic acid hybrid formed between said probe and said first target region of claim 3.

9. A composition comprising a nucleic acid hybrid formed between said probe and said first target region of claim 4.

10. A probe mix comprising the probe of claim 1 and at least one helper probe having from 12 to 100 bases which is able to hybridize to a second target nucleic acid region present in the nucleic acid containing said first target region under said conditions.

11. The probe mix of claim 10, wherein said second target region has a sequence selected from the group consisting of SEQ ID NO. 13, SEQ ID NO. 14, SEQ ID NO. 17, SEQ ID NO. 18, SEQ ID NO. 35, SEQ ID NO. 36, SEQ ID NO. 37 and SEQ ID NO. 38.

12. The probe mix of claim 11, wherein the base sequence of said helper probe is at least 80% complementary to the base sequence of said second target region.

13. The probe mix of claim 12, wherein the base sequence of said base sequence region is at least 80% complementary to the base sequence of said first target region.

14. The probe mix of claim 12, wherein the base sequence of said base sequence region is fully complementary to the base sequence of said first target region

15. The probe mix of claim 12, wherein the base sequence of said probe is at least 80% complementary to the base sequence of said first target region.

16. The probe mix of claim 12, wherein said at least one helper probe includes first and second helper probes, wherein the base sequence of said second target region for said first helper probe is selected from the group consisting of SEQ ID NO. 13, SEQ ID NO. 17, SEQ ID NO. 35 and SEQ ID NO. 37, and wherein the base sequence of said second target region for said second helper probe is selected from the group consisting of SEQ ID NO. 14, SEQ ID NO. 18, SEQ ID NO. 36 and SEQ ID NO. 38.

17. The probe mix of claim 16, wherein the base sequence of said base sequence region is at least 80% complementary to the base sequence of said first target region.

18. The probe mix of claim 16, wherein the base sequence of said base sequence region is fully complementary to the base sequence of said first target region

19. The probe mix of claim 16, wherein the base sequence of said probe is at least 80% complementary to the base sequence of said first target region.

31. The kit of claim 32, wherein the base sequence of said probe is at least 80% homologous to the base sequence of said first sequence.

32. A method for detecting the presence of Neisseria meningitidis subtypes A, C and L in a sample, said method comprising the steps of:

(a) contacting said sample with said probe of claim 1 under stringent hybridization assay conditions; and

(b) detecting the presence of said probe as an indication of the presence of at least one of Neisseria meningtidis subtypes A, C and L in said sample.

33. The method of claim 32, wherein the base sequence of said base sequence region is at least 80% complementary to the base sequence of said first target region.

34. The method of claim 32, wherein the base sequence of said base sequence region is fully complementary to the base sequence of said first target region.

35. The method of claim 32, wherein the base sequence of said probe is at least 80% complementary to the base sequence of said first target region.

36. The method of claim 32 further comprising the step of providing to said sample at least one amplification oligonucleotide able to bind to nucleic acid from Neisseria meningitidis under amplification conditions prior to said detecting step.

37. The method of claim 36, wherein said amplification oligonucleotide comprises a first base sequence which is at least 80% homologous to the base sequence of a second sequence selected from the group consisting of SEQ ID NO. 7, SEQ ID NO. 9, SEQ ID NO. 43 and SEQ ID NO. 45, wherein the base sequence of said amplification oligonucleotide consists of said first base sequence and, optionally, a 5′

sequence which is recognized by an RNA polymerase or which enhances initiation or elongation by an RNA polymerase.

38. The method of claim 37, wherein the base sequence of said base sequence region is at least 80% complementary to the base sequence of said first target region.

39. The method of claim 37, wherein the base sequence of said base sequence region is fully complementary to the base sequence of said first target region.

40. The method of claim 37, wherein the base sequence of said probe is at least 80% complementary to the base sequence of said first target region.

41. The method of claim 40, wherein said at least one amplification oligonucleotide includes first and second amplification oligonucleotides, wherein the base sequence of said second sequence for said first amplification oligonucleotide is SEQ ID NO. 7 or SEQ ID NO. 43, and wherein the base sequence of said second sequence for said second amplification oligonucleotide is SEQ ID NO. 9 or SEQ ID NO. 45.

20. A kit for use in detecting the presence of Neisseria meningitidis in a sample, said kit comprising:

a hybridization assay probe comprising a base sequence region which is able to form a detectable hybrid with nucleic acid from Neisseria meningitidis subtypes A, C and L and which is unable to form a detectable hybrid with nucleic acid from Neisseria gonorrhoeae under stringent hybridization assay conditions, wherein the base sequence of said region is at least 80% homologous to the base sequence of a first sequence selected from the group consisting of SEQ ID NO. 11, SEQ ID NO. 15, SEQ ID NO. 25 and SEQ ID NO. 27, and wherein said probe does not comprise any other base sequence region which is able to form a detectable hybrid with nucleic acid from Neisseria meningitidis subtype A, C or L or Neisseria gonorrhoeae under said conditions; and

at least one helper probe which is able to hybridize to nucleic acid from Neisseria meningitidis under stringent hybridization assay conditions, wherein the base sequence of said helper probe is at least 80% homologous to a the base sequence of a second sequence selected from the group consisting of SEQ ID NO. 13, SEQ ID NO. 14, SEQ ID NO. 17, SEQ ID NO. 18, SEQ ID NO. 35, SEQ ID NO. 36, SEQ ID NO. 37 and SEQ ID NO. 38.

21. The kit of claim 20, wherein the base sequence of said region is fully homologous to the base sequence of said first sequence.

22. The kit of claim 20, wherein the base sequence of said probe is at least 80% homologous to the base sequence of said first sequence.

23. The kit of claim 20, wherein said at least one helper probe includes first and second helper probes, wherein the base sequence of said second sequence for said first helper probe is selected from the group consisting of SEQ ID NO. 13, SEQ ID NO. 17, SEQ ID NO. 35 and SEQ ID NO. 37, and wherein the base sequence of said second sequence for said second helper probe is selected from the group consisting of SEQ ID NO. 14, SEQ ID NO. 18, SEQ ID NO. 36 and SEQ ID NO. 38.

24. The kit of claim 23, wherein the base sequence of said region is fully homologous to the base sequence of said first sequence.

25. The kit of claim 23, wherein the base sequence of said probe is at least 80% homologous to the base sequence of said first sequence.

26. A kit for use in detecting the presence of Neisseria meningitidis in a sample, said kit comprising:

a hybridization assay probe comprising a base sequence region which is able to form a detectable hybrid with nucleic acid from Neisseria meningitidis subtypes A, C and L and which is unable to form a detectable hybrid with nucleic acid from Neisseria gonorrhoeae under stringent hybridization assay conditions, wherein the base sequence of said region is at least 80% homologous to the base sequence of a first sequence selected from the group consisting of SEQ ID NO. 11, SEQ ID NO. 15, SEQ ID NO. 25 and SEQ ID NO. 27, and wherein said probe does not comprise any other base sequence region which is able to form a detectable hybrid with nucleic acid from Neisseria meningitidis subtype A, C or L or Neisseria gonorrhoeae under said conditions; and

at least one amplification oligonucleotide which is able to bind to nucleic acid from Neisseria meningitidis under amplification conditions, said amplification oligonucleotide comprising a first base sequence which is at least 80% homologous to the base sequence of a second sequence selected from the group consisting of SEQ ID NO. 7, SEQ ID NO. 9, SEQ ID NO. 43 and SEQ ID NO. 45, wherein the base sequence of said amplification oligonucleotide consists of said first base sequence and, optionally, a 5′

sequence which is recognized by an RNA polymerase or which enhances initiation or elongation by an RNA polymerase.

27. The kit of claim 26, wherein the base sequence of said region is fully homologous to the base sequence of said first sequence.

28. The kit of claim 27, wherein the base sequence of said probe is at least 80% homologous to the base sequence of said first sequence.

29. The kit of claim 26, wherein said at least one amplification oligonucleotide includes first and second amplification oligonucleotides, wherein the base sequence of said second sequence for said first amplification oligonucleotide is SEQ ID NO. 7 or SEQ ID NO. 43, and wherein the base sequence of said second sequence for said second amplification oligonucleotide is SEQ ID NO. 9 or SEQ ID NO. 45.

30. The kit of claim 29, wherein the base sequence of said region is fully homologous to the base sequence of said first sequence.