Detection of human papillomavirus
View Patent ↗There is provided an in vitro method of detecting human papillomavirus nucleic acid in a sample, comprising: (a) contacting said sample with forward and reverse oligonucleotide primers, wherein said primers bind to target sites in the human papillomavirus L1 gene, or the complement thereof, under conditions suitable to promote amplification of a portion of said human papillomavirus L1 gene or complement, thereby generating an amplicon; (b) contacting said amplicon with a probe, wherein the probe binds to a target site within said amplicon; and (c) detecting binding of said probe to said amplicon; wherein said forward primer binds to a target site having the sequence SEQ ID NO: 1; and wherein said reverse primer binds to a target site having the sequence SEQ ID NO: 2.
1. An in vitro method of detecting human papillomavirus nucleic acid in a sample, comprising:
contacting said sample with forward and reverse oligonucleotide primers, wherein said primers bind to target sites in the human papillomavirus L1 gene, or the complement thereof, under conditions suitable to promote amplification of a portion of said human papillomavirus L1 gene or complement, thereby generating an amplicon;
wherein said forward primer binds to a target site having the sequence SEQ ID NO: 1, and
wherein said reverse primer binds to a target site having the sequence SEQ ID NO: 2;
contacting said amplicon with a set of probes, wherein said set of probes comprises
(i) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 15;
(ii) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 16;
(iii) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 17; and
(iv) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 18; and
detecting binding of said set of probes to said amplicon.
2. The method of claim 1 , wherein said set of probes comprises:
(i) a probe comprising a nucleic acid having the sequence SEQ ID NO: 15,
(ii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 16,
(iii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 17, and
(iv) a probe comprising a nucleic acid having the sequence SEQ ID NO: 18.
3. The method of claim 1 , wherein each probe of said set of probes is immobilised onto a different solid support or platform.
4. The method of claim 3 , wherein said different solid supports or platforms are uniquely identifiable beads or microspheres.
5. The method of claim 4 , wherein following detecting binding of said amplicon to said set of probes, said method further comprises sorting said beads or microspheres according to their unique identification.
6. The method of claim 1 , wherein said forward primer comprises a nucleic acid having the sequence SEQ ID NO: 3.
7. The method of claim 6 , wherein said forward primer comprises a nucleic acid having at least 85% sequence identity to a sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 or SEQ ID NO: 8.
8. The method of claim 7 wherein said forward primer consists of a nucleic acid having a sequence selected from SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 or SEQ ID NO: 8.
9. The method of claim 1 , wherein said reverse primer comprises a nucleic acid having the sequence SEQ ID NO: 9.
10. The method of claim 9 , wherein said reverse primer comprises a nucleic acid having at least 85% sequence identity to a sequence selected from SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13 or SEQ ID NO: 14.
11. The method of claim 1 , wherein said set of probes further comprises at least one additional probe comprising a nucleic acid having at least 85% sequence identity to a sequence selected from the group consisting of: SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33 and SEQ ID NO: 34.
12. An in vitro method of detecting human papillomavirus nucleic acid in a sample, comprising:
contacting said sample with forward and reverse oligonucleotide primers, wherein said primers bind to target sites in the human papillomavirus L1 gene, or the complement thereof, under conditions suitable to promote amplification of a portion of said human papillomavirus L1 gene or complement, thereby generating an amplicon;
contacting said amplicon with a set of probes;
wherein said set of probes comprises
(i) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 15;
(ii) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 16;
(iii) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 17; and
(iv) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 18; and
detecting binding of said set of probes to said amplicon.
13. The method of claim 12 , wherein said set of probes further comprises at least one additional probe comprising a nucleic acid having at least 85% sequence identity to a sequence selected from the group consisting of: SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33 and SEQ ID NO: 34.
14. The method of claim 12 , wherein each probe of said set of probes is immobilised onto a different solid support or platform.
15. The method of claim 14 , wherein said different solid supports or platforms are uniquely identifiable beads or microspheres.
16. The method of claim 15 , wherein following detecting binding of said amplicon to said set of probes, said method further comprises sorting said beads or microspheres according to their unique identification.
17. The method of claim 12 , wherein said set of probes comprises:
(i) a probe comprising a nucleic acid having the sequence SEQ ID NO: 15,
(ii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 16,
(iii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 17, and
(iv) a probe comprising a nucleic acid having the sequence SEQ ID NO: 18.
18. An in vitro method of detecting human papillomavirus nucleic acid in a sample, comprising:
contacting said sample with a set of probes that bind to a target site within the human papillomavirus L1 gene, or the complement thereof;
wherein said set of probes comprises
(i) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 15;
(ii) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 16;
(iii) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 17; and
(iv) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 18; and
detecting binding of said set of probes to said target site.
19. The method of claim 18 , wherein each probe of said set of probes is immobilised onto a different solid support or platform.
20. The method of claim 19 , wherein said different solid supports or platforms are uniquely identifiable beads or microspheres.
21. The method of claim 20 , wherein following binding of said set of probes to said target site, said method further comprises sorting said beads or microspheres according to their unique identification.
22. The method of claim 18 , wherein said set of probes further comprises at least one additional probe comprising a nucleic acid having at least 85% sequence identity to a sequence selected from the group consisting of: SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33 and SEQ ID NO: 34.
23. The method of claim 18 , wherein said set of probes comprises:
(i) a probe comprising a nucleic acid having the sequence SEQ ID NO: 15,
(ii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 16,
(iii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 17, and
(iv) a probe comprising a nucleic acid having the sequence SEQ ID NO: 18.
24. A set of probes that bind to a target sequence within the human papillomavirus L1 gene, or the complement thereof;
wherein said set of probes comprises:
(i) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 15;
(ii) a probe comprising a nucleic acid having at least 90% sequence identity to SEQ ID NO: 16;
(iii) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 17; and
(iv) a probe comprising a nucleic acid having at at least 90% sequence identity to SEQ ID NO: 18.
25. The set of probes of claim 24 , further comprising at least one additional probe comprising a nucleic acid having at least 85% sequence identity to a sequence selected from the group consisting of: SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33 and SEQ ID NO: 34.
26. The set of probes of claim 24 , wherein each probe of said set of probes is immobilised onto a different solid support or platform.
27. The set of probes of claim 26 , wherein said different solid supports or platforms are uniquely identifiable beads or microspheres.
28. The set of probes of claim 24 , comprising:
(i) a probe comprising a nucleic acid having the sequence SEQ ID NO: 15,
(ii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 16,
(iii) a probe comprising a nucleic acid having the sequence SEQ ID NO: 17, and
(iv) a probe comprising a nucleic acid having the sequence SEQ ID NO: 18.