Detection of nucleic acids from multiple types of human papillomavirus
View Patent ↗Nucleic acid oligonucleotide sequences are disclosed which include amplification oligomers and probe oligomers which are useful for detecting multiple types of human papillomaviruses (HPV) associated with cervical cancer. Methods for detecting multiple HPV types in biological specimens by amplifying HPV nucleic acid sequences in vitro and detecting the amplified products are disclosed.
1. A mixture of oligomers in which individual oligomer sequences are selected from the group consisting of SEQ ID NO:11 to SEQ ID NO:17, SEQ ID NO:44 to SEQ ID NO:54 and SEQ ID NO:58, which includes the complementary oligomer sequences or RNA equivalents of the specified sequences, wherein the mixture includes:
SEQ ID Nos. 11 to 17, or complementary oligomer sequences or RNA equivalents of the oligomer sequences;
SEQ ID Nos. 11 to 15 and 17, or complementary oligomer sequences or RNA equivalents of the oligomer sequences;
SEQ ID Nos. 11 to 15 and 17 and at least one oligomer of SEQ ID NO:44 to SEQ ID NO:54 and SEQ ID NO:58, or complementary oligomer sequences or RNA equivalents of the oligomer sequences; or
SEQ ID Nos. 11, 12, 14, 15, 17, 44, 45 and 52, or complementary oligomer sequences or RNA equivalents of the oligomer sequences
wherein at least one of the oligomer sequences in the mixture of oligomers includes a label joined directly or indirectly to the oligomer sequence.
2. The mixture of oligomers of claim 1 , wherein each oligomer sequence in the mixture of oligomers includes a label joined directly or indirectly to the oligomer sequence.
3. The mixture of oligomers of claim 1 , wherein each oligomer sequence in the mixture of oligomers includes a label that is a chemiluminescent compound.
4. The mixture of oligomers of claim 1 , wherein each oligomer sequence in the mixture of oligomers has a backbone comprising at least one 2′-methoxy RNA group.
5. The mixture of oligomers of claim 1 contained in a kit.
6. A method of detecting human papillomavirus (HPV) nucleic acid present in a biological sample, comprising the steps of:
contacting nucleic acid in a biological sample containing RNA of at least one of HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68 with a mixture of amplification oligomers that amplify a HPV sequence in an E6/E7 target region sequence, in which the mixture is made up of
first amplification oligomers of SEQ ID Nos. 19 or 42, 21, 23, 25, 27, 29, 31, 33 and 35, and second amplification oligomers of SEQ ID Nos. 37, 38, 39, 40 and 41, or complementary oligomer sequences or RNA equivalents of the oligomer sequences;
first amplification oligomers of SEQ ID Nos. 18, 20, 22, 24, 26, 28, 30, 32, 34 and 36, and second amplification oligomers of SEQ ID Nos. 37, 38, 39, 40 and 41, or complementary oligomer sequences or RNA equivalents of the oligomer sequences;
first amplification oligomers of SEQ ID Nos. 19 or 42, 21, 23, 25, 27, 29, 31, 33 and 35, and second amplification oligomers of SEQ ID Nos. 38, 39, 40 and 41, or complementary oligomer sequences or RNA equivalents of the oligomer sequences; or
first amplification oligomers of SEQ ID Nos. 18, 20, 22, 24, 26, 28, 30, 32, 34 and 36, and second amplification oligomers of SEQ ID Nos. 38, 39, 40 and 41, or complementary oligomer sequences or RNA equivalents of the oligomer sequences;
amplifying a HPV sequence from the target region sequence in at least one HPV type by using the amplification oligomers and a nucleic acid polymerase in vitro to produce an HPV amplified product; and
detecting the amplified product by using the mixture of oligomers of claim 1 .
7. The method of claim 6 , further comprising a step of separating RNA of at least one of HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68 from other components in the sample by contacting HPV RNA in the sample with a capture oligomer and separating a complex that includes the capture oligomer and HPV RNA from other components in the sample before the amplifying step.
8. The method of claim 7 , wherein the capture oligomer is present in a mixture of capture oligomers made up of at least two oligomers in which individual oligomer sequences are selected from the group consisting of SEQ ID Nos. 1 to 10, which includes the complementary oligomer sequences or RNA equivalents of the specified sequences.
9. The method of claim 7 , wherein the capture oligomer is present in a mixture of capture oligomers made up of:
at least two oligomers selected from SEQ ID Nos. 2, 4, 6, 8 and 10 with a ligand moiety joined to each oligomer;
oligomers of SEQ ID Nos. 2, 4, 6, 8 and 10 with a ligand moiety joined to each oligomer;
at least two oligomers selected from SEQ ID Nos. 1, 3, 5, 7 and 9; or oligomers of SEQ ID Nos. 1, 3, 5, 7 and 9.
10. The method of claim 6 , wherein the amplifying step uses an amplification process that is substantially isothermal.
11. The method of claim 6 , wherein the amplifying step uses a transcription-associated amplification method.
12. The method of claim 6 , wherein the contacting step further includes introducing a non-HPV internal control sequence into the sample,
the amplifying step further includes amplifying the non-HPV internal control sequence to produce an amplified internal control sequence, and
the detecting step further includes detecting the amplified internal control sequence to produce a signal that indicates that the method steps have been performed appropriately.
13. A method of detecting human papillomavirus (HPV) nucleic acid present in a biological sample, comprising the steps of:
contacting nucleic acid in a biological sample containing RNA of at least one of HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68 with a mixture of amplification oligomers that amplify a HPV sequence in an E6/E7 target region sequence;
amplifying a HPV sequence from the target region sequence in at least one HPV type by using the amplification oligomers and a nucleic acid polymerase in vitro to produce an HPV amplified product; and
detecting the amplified product by using at the mixture of oligomers of claim 1 .
14. The method of claim 13 , further comprising a step of separating RNA of at least one of HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68 from other components in the sample by contacting HPV RNA in the sample with a capture oligomer and separating a complex that includes the capture oligomer and HPV RNA from other components in the sample before the amplifying step.
15. The method of claim 14 , wherein the capture oligomer is present in a mixture of capture oligomers made up of at least two oligomers in which individual oligomer sequences are selected from the group consisting of SEQ ID Nos. 1 to 10, which includes the complementary oligomer sequences or RNA equivalents of the specified sequences.
16. The method of claim 14 , wherein the capture oligomer is present in a mixture of capture oligomers made up of:
at least two oligomers selected from SEQ ID Nos. 2, 4, 6, 8 and 10 with a ligand moiety joined to each oligomer;
oligomers of SEQ ID Nos. 2, 4, 6, 8 and 10 with a ligand moiety joined to each oligomer;
at least two oligomers selected from SEQ ID Nos. 1, 3, 5, 7 and 9; or
oligomers of SEQ ID Nos. 1, 3, 5, 7 and 9.
17. The method of claim 13 , wherein the amplifying step uses an amplification process that is substantially isothermal.
18. The method of claim 13 , wherein the amplifying step uses a transcription-associated amplification method.
19. The method of claim 13 , wherein the contacting step further includes introducing a non-HPV internal control sequence into the sample,
the amplifying step further includes amplifying the non-HPV internal control sequence to produce an amplified internal control sequence, and
the detecting step further includes detecting the amplified internal control sequence to produce a signal that indicates that the method steps have been performed appropriately.