Kits to detect adenovirus nucleic acids
The disclosed invention is related to methods, compositions, kits and isolated nucleic acid sequences for targeting Adenovirus nucleic acid. Compositions include amplification oligomers and/or detection probe oligomers. Kits and methods comprise at least one of these oligomers.
1. A kit containing a detection probe configured to hybridize to a portion of an Adenovirus nucleic acid or to a portion of an amplicon generated therefrom, wherein the detection probe comprises a target hybridizing sequence selected from the group consisting of: SEQ ID NO:10, SEQ ID NO:17, SEQ ID NO:21, SEQ ID NO:29, and SEQ ID NO:36, and
wherein the detection probe further comprises one or more of
(a) a label,
(b) a substitution at the 2′ position of at least one ribose moiety, and
(c) a blocking moiety at or near the 3′ end of the detection probe, wherein the blocking moiety is configured to prevent enzyme-mediated extension of the detection probe in an amplification reaction.
2. The kit of claim 1 , wherein the detection probe further comprises the label.
3. The kit of claim 2 , wherein the label is a fluorescent or chemiluminescent label.
4. The kit of claim 3 , wherein the detection probe comprises the fluorescent label in combination with a quencher.
5. The kit of claim 1 , wherein the detection probe comprises the substitution at the 2′ position of at least one ribose moiety.
6. The kit of claim 5 , wherein the substitution is 2′-methoxy or 2′-halide.
7. The kit of claim 1 , wherein the detection probe further comprises the blocking moiety, wherein the blocking moiety is selected from the group consisting of an alkyl group, a non-nucleotide linker, an alkane-diol dideoxynucleotide residue, and cordycepin.
8. The kit of claim 1 , further comprising at least one amplification oligonucleotide.
9. A kit containing a detection probe configured to hybridize to a portion of an Adenovirus nucleic acid or to a portion of an amplicon generated therefrom, wherein the detection probe comprises a target hybridizing sequence selected from the group consisting of: SEQ ID NO:10, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:29, and SEQ ID NO:36,
wherein the detection probe further comprises one or more of
(a) a label,
(b) a substitution at the 2′ position of at least one ribose moiety, and
(c) a blocking moiety at or near the 3′ end of the detection probe, wherein the blocking moiety is configured to prevent enzyme-mediated extension of the detection probe in an amplification reaction, and
wherein the kit further comprises at least one amplification oligonucleotide, wherein:
(i) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:10, then the at least one amplification oligonucleotide is a forward primer comprising the target hybridizing sequence of SEQ ID NO:5 and a reverse primer comprising the target hybridizing sequence of SEQ ID NO:6 or SEQ ID NO:8;
(ii) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:17, then the at least one amplification oligonucleotide is a forward primer comprising the target hybridizing sequence of SEQ ID NO:11 or SEQ ID NO:12 and a reverse primer comprising the target hybridizing sequence of SEQ ID NO:13 or SEQ ID NO:15;
(iii) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:19, then the at least one amplification oligonucleotide is a forward primer comprising the target hybridizing sequence of SEQ ID NO:11 or SEQ ID NO:12 and a reverse primer comprising the target hybridizing sequence of SEQ ID NO:13 or SEQ ID NO:15;
(iv) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:21, then the at least one amplification oligonucleotide is two forward primers comprising the target hybridizing sequences of SEQ ID NO:25 and SEQ ID NO:26 and two reverse primers comprising the target hybridizing sequences of SEQ ID NO:27 and SEQ ID NO:28;
(v) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:23, then the at least one amplification oligonucleotide is two forward primers comprising the target hybridizing sequences of SEQ ID NO:25 and SEQ ID NO:26 and two reverse primers comprising the target hybridizing sequences of SEQ ID NO:27 and SEQ ID NO:28; or
(vi) if the detection probe comprises the target hybridizing sequence of SEQ ID NO:29, then the at least one amplification oligonucleotide is two forward primers comprising the target hybridizing sequences of SEQ ID NO:25 and SEQ ID NO:26 and two reverse primers comprising the target hybridizing sequences of SEQ ID NO:27 and SEQ ID NO:28.
10. A kit containing at least two detection probes configured to hybridize to a portion of an Adenovirus nucleic acid or to a portion of an amplicon generated therefrom, wherein each detection probe comprises a target hybridizing sequence selected from the group consisting of SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:36, and SEQ ID NO:37,
wherein each of the detection probes further comprises one or more of
(a) a label,
(b) a substitution at the 2′ position of at least one ribose moiety, and
(c) a blocking moiety at or near the 3′ end of the detection probe, wherein the blocking moiety is configured to prevent enzyme-mediated extension of the detection probe in an amplification reaction, and
wherein at least one of the at least two detection probes comprises the target hybridizing sequence of SEQ ID NO:21 or SEQ ID NO:36.
11. The kit of claim 10 , wherein each of the detection probes further comprises the label.
12. The kit of claim 11 , wherein the label is a fluorescent or chemiluminescent label.
13. The kit of claim 12 , wherein each detection probe comprises the fluorescent label in combination with a quencher.
14. The kit of claim 10 , wherein each detection probe further comprises the substitution at the 2′ position of at least one ribose moiety.
15. The kit of claim 14 , wherein the substitution is 2′-methoxy or 2′-halide.
16. The kit of claim 10 , wherein each detection probe further comprises the blocking moiety, wherein the blocking moiety is selected from the group consisting of an alkyl group, a non-nucleotide linker, an alkane-diol dideoxynucleotide residue, and cordycepin.
17. The kit of claim 10 , further comprising at least one amplification oligonucleotide.
18. The kit of claim 17 , wherein:
(i) if the at least two detection probes comprise the target hybridizing sequences of SEQ ID NO:21 and SEQ ID NO:23, then the at least one amplification oligonucleotide is a forward primer comprising the target hybridizing sequence of SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:31, SEQ ID NO:33, SEQ ID NO:34 and/or SEQ ID NO:35 and a reverse primer comprising the target hybridizing sequence of SEQ ID NO:27 and/or SEQ ID NO:28;
(ii) if the at least two detection probes comprise the target hybridizing sequences of SEQ ID NO:21 and SEQ ID NO:24, then the at least one amplification oligonucleotide is a forward primer comprising the target hybridizing sequence of SEQ ID NO:12 and a reverse primer comprising the target hybridizing sequence of SEQ ID NO:15; or
(iii) if the at least two detection probes comprise the target hybridizing sequences of SEQ ID NO:36 and SEQ ID NO:37, then the at least one amplification oligonucleotide is two forward primers comprising the target hybridizing sequences of SEQ ID NO:25 and SEQ ID NO:26 and two reverse primers comprising the target hybridizing sequences of SEQ ID NO:27 and SEQ ID NO:28.