Compositions and methods for detecting Zika virus nucleic acid
Disclosed are nucleic acid oligomers, including amplification oligomers, capture probes, and detection probes, for detection of Zika virus nucleic acid. Also disclosed are methods of specific nucleic acid amplification and detection using the disclosed oligomers, as well as corresponding reaction mixtures and kits.
1. A combination of at least two amplification oligomers for amplifying a Zika virus nucleic acid in a sample, the oligomer combination comprising:
at least two amplification oligomers configured to amplify a target sequence corresponding to a Zika virus target nucleic acid, wherein the target sequence consists of SEQ ID NO: 189 or SEQ ID NO: 190, the RNA equivalent of SEQ ID NO: 189 or SEQ ID NO: 190, or the DNA or RNA complement of SEQ ID NO: 189 or SEQ ID NO: 190,
wherein the first amplification oligomer and the second amplification oligomer are configured to hybridize to opposite ends of the target sequence to generate amplification products;
wherein the first amplification oligomer comprises (i) a target-hybridizing sequence consisting of from 17 to 26 contiguous nucleobases of SEQ ID NO: 24, wherein said contiguous nucleobases of the target-hybridizing sequence include SEQ ID NO: 25, or (ii) a target-hybridizing sequence consisting of SEQ ID NO: 27, and
wherein the second amplification oligomer comprises a target-hybridizing sequence that is selected from the group consisting of SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 126, and SEQ ID NO: 128, wherein the second amplification oligomer target-hybridizing sequence is joined at its 5′ end to a T7 promoter sequence.
2. An amplification reaction mixture, comprising the at least two amplification oligomers of claim 1 , wherein (i) the mixture is a buffered aqueous solution; or (ii) the mixture is a dried composition.
3. A kit comprising the at least two amplification oligomers of claim 1 .
4. The combination of claim 1 , wherein the T7 promoter sequence consists of SEQ ID NO:179.
5. The combination of claim 1 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:113 and SEQ ID NO:126.
6. The combination of claim 5 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, and SEQ ID NO:27.
7. The combination of claim 6 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:74 and SEQ ID NO:76.
8. The combination of claim 1 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:128 and SEQ ID NO:115.
9. The combination of claim 8 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, and SEQ ID NO:23.
10. The combination of claim 8 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:80 and SEQ ID NO:87.
11. The combination of claim 1 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:16 and SEQ ID NO:80,
(ii) SEQ ID NO:16 and SEQ ID NO:74,
(iii) SEQ ID NO:11 and SEQ ID NO:80,
(iv) SEQ ID NO:11 and SEQ ID NO:74,
(v) SEQ ID NO:12 and SEQ ID NO:87,
(vi) SEQ ID NO:12 and SEQ ID NO:74,
(vii) SEQ ID NO:17 and SEQ ID NO:87,
(viii) SEQ ID NO:17 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80, and
(x) SEQ ID NO:18 and SEQ ID NO:74.
12. The combination of claim 1 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:12 and SEQ ID NO:87,
(ii) SEQ ID NO:13 and SEQ ID NO:87,
(iii) SEQ ID NO:14 and SEQ ID NO:74,
(iv) SEQ ID NO:15 and SEQ ID NO:80,
(v) SEQ ID NO:15 and SEQ ID NO:87,
(vi) SEQ ID NO:16 and SEQ ID NO:80,
(vii) SEQ ID NO:16 and SEQ ID NO:87,
(viii) SEQ ID NO:18 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80,
(x) SEQ ID NO:20 and SEQ ID NO:74,
(xi) SEQ ID NO:21 and SEQ ID NO:74,
(xii) SEQ ID NO:21 and SEQ ID NO:87,
(xiii) SEQ ID NO:22 and SEQ ID NO:80, and
(xiv) SEQ ID NO:23 and SEQ ID NO:74.
13. A method for determining the presence or absence of a Zika virus nucleic acid in a sample, the method comprising the steps of:
(A) contacting a sample with a combination of at least two amplification oligomers configured to amplify a target sequence corresponding to a Zika virus target nucleic acid, wherein the target sequence consists of SEQ ID NO: 189 or SEQ ID NO: 190, the RNA equivalent of SEQ ID NO: 189 or SEQ ID NO: 190, or the DNA or RNA complement of SEQ ID NO: 189 or SEQ ID NO: 190,
wherein the first amplification oligomer and the second amplification oligomer are configured to hybridize to opposite ends of the target sequence to generate amplification products;
wherein the first amplification oligomer comprises (i) a target-hybridizing sequence consisting of from 17 to 26 contiguous nucleobases of SEQ ID NO: 24, wherein said contiguous nucleobases of the target-hybridizing sequence include SEQ ID NO: 25, or (ii) a target-hybridizing sequence consisting of SEQ ID NO: 27, and
wherein the second amplification oligomer comprises a target-hybridizing sequence that is selected from the group consisting of SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 126, and SEQ ID NO: 128, wherein the second amplification oligomer target-hybridizing sequence is joined at its 5′ end to a T7 promoter sequence;
(B) performing an in vitro nucleic acid amplification reaction, wherein any Zika virus target nucleic acid present in the sample is used as a template for generating an amplification product; and
(C) detecting the presence or absence of the amplification product, thereby determining the presence or absence of the Zika virus nucleic acid in the sample.
14. The method of claim 13 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:113 and SEQ ID NO:126.
15. The method of claim 14 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23 and SEQ ID NO: 27.
16. The method of claim 15 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:74 and SEQ ID NO:76.
17. The method of claim 13 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:128 and SEQ ID NO:115.
18. The method of claim 17 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, and SEQ ID NO:23.
19. The method of claim 17 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:80 and SEQ ID NO:87.
20. The method of claim 13 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:16 and SEQ ID NO:80,
(ii) SEQ ID NO:16 and SEQ ID NO:74,
(iii) SEQ ID NO:11 and SEQ ID NO:80,
(iv) SEQ ID NO:11 and SEQ ID NO:74,
(v) SEQ ID NO:12 and SEQ ID NO:87,
(vi) SEQ ID NO:12 and SEQ ID NO:74,
(vii) SEQ ID NO:17 and SEQ ID NO:87,
(viii) SEQ ID NO:17 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80, and
(x) SEQ ID NO:18 and SEQ ID NO:74.
21. The method of claim 13 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:12 and SEQ ID NO:87,
(ii) SEQ ID NO:13 and SEQ ID NO:87,
(iii) SEQ ID NO:14 and SEQ ID NO:74,
(iv) SEQ ID NO:15 and SEQ ID NO:80,
(v) SEQ ID NO:15 and SEQ ID NO:87,
(vi) SEQ ID NO:16 and SEQ ID NO:80,
(vii) SEQ ID NO:16 and SEQ ID NO:87,
(viii) SEQ ID NO:18 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80,
(x) SEQ ID NO:20 and SEQ ID NO:74,
(xi) SEQ ID NO:21 and SEQ ID NO:74,
(xii) SEQ ID NO:21 and SEQ ID NO:87,
(xiii) SEQ ID NO:22 and SEQ ID NO:80, and
(xiv) SEQ ID NO:23 and SEQ ID NO:74.
22. A multiplex method for determining the presence or absence of a Zika virus nucleic acid in a sample, the method comprising the steps of:
(A) contacting a sample with at least two combinations of at least two amplification oligomers, each configured to amplify a separate target sequence of a Zika virus target nucleic acid, wherein the at least two combinations of at least two amplification oligomers comprise (i) a first combination comprising the at least two amplification oligomers of claim 1 combined with (ii) a second combination of at least two amplification oligomers;
(B) performing an in vitro nucleic acid amplification reaction, wherein the at least two amplification oligomers of (i) and/or (ii) hybridize their respective target sequence present in the sample to generate an amplification product(s); and
(C) detecting the presence or absence of the amplification product(s), thereby determining the presence or absence of the Zika virus nucleic acid in the sample.
23. The multiplex method of claim 22 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:113 and SEQ ID NO:126.
24. The multiplex method of claim 23 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23 and SEQ ID NO: 27.
25. The multiplex method of claim 24 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:74 and SEQ ID NO:76.
26. The multiplex method of claim 22 , wherein the second amplification oligomer target-hybridizing sequence is selected from the group consisting of SEQ ID NO:128 and SEQ ID NO:115.
27. The multiplex method of claim 26 , wherein the nucleotide sequence of the first amplification oligomer is selected from the group consisting of SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, and SEQ ID NO:23.
28. The multiplex method of claim 26 , wherein the nucleotide sequence of second amplification oligomer is selected from the group consisting of SEQ ID NO:80 and SEQ ID NO:87.
29. The multiplex method of claim 22 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:16 and SEQ ID NO:80,
(ii) SEQ ID NO:16 and SEQ ID NO:74,
(iii) SEQ ID NO:11 and SEQ ID NO:80,
(iv) SEQ ID NO:11 and SEQ ID NO:74,
(v) SEQ ID NO:12 and SEQ ID NO:87,
(vi) SEQ ID NO:12 and SEQ ID NO:74,
(vii) SEQ ID NO:17 and SEQ ID NO:87,
(viii) SEQ ID NO:17 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80, and
(x) SEQ ID NO:18 and SEQ ID NO:74.
30. The multiplex method of claim 22 , wherein the nucleotide sequences of the first amplification oligomer and second amplification oligomer are selected from the group consisting of
(i) SEQ ID NO:12 and SEQ ID NO:87,
(ii) SEQ ID NO:13 and SEQ ID NO:87,
(iii) SEQ ID NO:14 and SEQ ID NO:74,
(iv) SEQ ID NO:15 and SEQ ID NO:80,
(v) SEQ ID NO:15 and SEQ ID NO:87,
(vi) SEQ ID NO:16 and SEQ ID NO:80,
(vii) SEQ ID NO:16 and SEQ ID NO:87,
(viii) SEQ ID NO:18 and SEQ ID NO:74,
(ix) SEQ ID NO:18 and SEQ ID NO:80,
(x) SEQ ID NO:20 and SEQ ID NO:74,
(xi) SEQ ID NO:21 and SEQ ID NO:74,
(xii) SEQ ID NO:21 and SEQ ID NO:87,
(xiii) SEQ ID NO:22 and SEQ ID NO:80, and
(xiv) SEQ ID NO:23 and SEQ ID NO:74.