Compositions and methods for detecting human papillomavirus nucleic acid
Disclosed are nucleic acid oligomers, including amplification oligomers, capture probes, and detection probes, for detection of a human papillomavirus (HPV) 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 target capture reaction mixture for selectively separating a human papillomavirus type 33 (HPV33) and/or type 31 (HPV31) target nucleic acid from other components in a sample, wherein the target capture reaction mixture comprises
(a) an HPV33-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:50; and
(b) an HPV31-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:52;
wherein each of the HPV33-specific and HPV31-specific capture probe oligomers further comprises an immobilized probe-binding region that is not complementary to the HPV33 or HPV31 target nucleic acid.
2. The capture probe oligomer of claim 1 , wherein the immobilized probe-binding region comprises a homopolymeric tail of about 10 to 40 nucleotides.
3. The capture probe oligomer of claim 2 , wherein the immobilized probe-binding region is selected from the group consisting of A 14 to A 40 and T 3 A 14 to T 3 A 30 .
4. The capture probe oligomer of claim 3 , wherein
the nucleotide sequence of the HPV33-specific capture probe oligomer consists of SEQ ID NO:51, and/or
the nucleotide sequence of the HPV31-specific capture probe oligomer consists of SEQ ID NO:53.
5. A kit for selectively separating a human papillomavirus type 33 (HPV33) and/or type 31 (HPV31) target nucleic acid from other components in a sample, wherein the kit comprises
(a) an HPV33-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:50; and
(b) an HPV31-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:52;
wherein each of the HPV33-specific and HPV31-specific capture probe oligomers further comprises an immobilized probe-binding region that is not complementary to the HPV33 or HPV31 target nucleic acid.
6. The kit of claim 5 , wherein the immobilized probe-binding region comprises a homopolymeric tail of about 10 to 40 nucleotides.
7. The kit of claim 6 , wherein the immobilized probe-binding region is selected from the group consisting of A 14 to A 40 and T 3 A 14 to T 3 A 30 .
8. The kit of claim 7 , wherein
the nucleotide sequence of the HPV33-specific capture probe oligomer consists of SEQ ID NO:51, and/or
the nucleotide sequence of the HPV31-specific capture probe oligomer consists of SEQ ID NO:53.
9. The kit of claim 5 , wherein the kit is for amplifying and/or detecting the HPV33 and/or HPV31 target nucleic acid, and wherein the kit further comprises at least one of
(i) an amplification oligomer combination for amplification of the HPV33 and/or HPV31 target nucleic acid; and
(ii) a detection probe oligomer for determining the presence or absence of an HPV33 and/or HPV31 amplification product.
10. A multiplex method for selectively separating a human papillomavirus type 33 (HPV33) and/or type 31 (HPV31) target nucleic acid from other components in a sample and detecting the separated target nucleic acid, said method comprising:
(1) contacting a sample, said sample suspected of containing HPV33 and/or HPV31, with
(a) an HPV33-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:50; and
(b) an HPV31-specific capture probe oligomer comprising a target-hybridizing sequence consisting of SEQ ID NO:52;
wherein each of the HPV33-specific and HPV31-specific capture probe oligomers further comprises an immobilized probe-binding region that is not complementary to the HPV33 or HPV31 target nucleic acid;
wherein the target-hybridizing sequence of the HPV33-specific and/or HPV31-specific capture probe oligomer specifically hybridizes to a target sequence in any HPV33 and/or HPV31 target nucleic acid, if present in the sample, so as to form a target-sequence:capture-probe complex;
(2) contacting any target-sequence:capture-probe complex from step (1) with an immobilized probe attached to a solid support, wherein the immobilized probe-binding region of the HPV33-specific and/or HPV31-specific capture probe in said complex hybridizes to the immobilized probe;
(3) separating the solid support and any hybridized target-sequence:capture-probe complex from other sample components, thereby selectively separating the HPV33 and/or HPV31 target nucleic acid from the other sample components;
(4) contacting any separated target nucleic acid with an amplification oligomer combination for specifically amplifying an HPV33 and/or HPV31 nucleic acid target region;
(5) performing an in vitro nucleic acid amplification reaction, wherein any separated HPV33 and/or HPV31 target nucleic acid, if present, is used as a template for generating an HPV33 and/or HPV31 amplification product; and
(6) detecting the presence or absence of the HPV33 and/or HPV31 amplification product.
11. The method of claim 10 , wherein the immobilized probe-binding region comprises a homopolymeric tail of about 10 to 40 nucleotides.
12. The method of claim 11 , wherein the immobilized probe-binding region is selected from the group consisting of A 14 to A 40 and T 3 A 14 to T 3 A 30 .
13. The method of claim 10 , wherein the solid support is a paramagnetic bead.
14. The method of claim 10 , wherein
the HPV33 nucleic acid target region to be amplified comprises a sequence contained within SEQ ID NO:83 from about nucleotide position 128 to about nucleotide position 164; and/or
the HPV31 nucleic acid target region to be amplified comprises a sequence contained within SEQ ID NO:84 from about nucleotide position 675 to about nucleotide position 735.