COMPOSITIONS FOR DETECTING BORDETELLA PARAPERTUSSIS NUCLEIC ACID
Disclosed are nucleic acid oligomers, including amplification oligomers and detection probes, for detection of Bordetella pertussis and Bordetella parapertussis 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 composition for determining the presence or absence of Bordetella parapertussis (Bpp) in a sample, said composition comprising:
an amplification oligomer combination comprising first and second Bpp-specific amplification oligomers capable of amplifying a target region of a Bpp target nucleic acid, wherein the first and second Bpp-specific amplification oligomers comprise, respectively, a first Bpp-specific target-hybridizing sequence and a second Bpp-specific target-hybridizing sequence, wherein
(a) the first Bpp-specific target-hybridizing sequence is SEQ ID NO:30, SEQ ID NO:36, or SEQ ID NO:39, or an RNA equivalent or DNA/RNA chimeric thereof, and
the second Bpp-specific target-hybridizing sequence is SEQ ID NO:32, SEQ ID NO:38, or SEQ ID NO:41, or an RNA equivalent or DNA/RNA chimeric thereof;
(b) the first Bpp-specific target-hybridizing sequence is SEQ ID NO:33, or an RNA equivalent or DNA/RNA chimeric thereof, and
the second Bpp-specific target-hybridizing sequence is SEQ ID NO:35, or an RNA equivalent or DNA/RNA chimeric thereof; or
(c) the first Bpp-specific target-hybridizing sequence is SEQ ID NO:42, or an RNA equivalent or DNA/RNA chimeric thereof, and
the second Bpp-specific target-hybridizing sequence is SEQ ID NO:44, or an RNA equivalent or DNA/RNA chimeric thereof; and
a Bpp-specific detection probe oligomer comprising a Bpp-specific detection probe target-hybridizing sequence that is from about 15 to about 35 nucleotides in length and is configured to hybridize to a target sequence contained within a Bpp amplicon amplifiable by the first and second Bpp-specific amplification oligomers, wherein the Bpp-specific detection probe oligomer comprises a fluorescent or chemiluminescent label.
2 . The composition of claim 1 , wherein the first and second Bpp specific target-hybridizing sequences are, respectively, selected from the group consisting of
(i) SEQ ID NO:30, or an RNA equivalent or DNA/RNA chimeric thereof, and SEQ ID NO:32, or an RNA equivalent or DNA/RNA chimeric thereof;
(ii) SEQ ID NO:33, or an RNA equivalent or DNA/RNA chimeric thereof, and SEQ ID NO:35, or an RNA equivalent or DNA/RNA chimeric thereof;
(iii) SEQ ID NO:36, or an RNA equivalent or DNA/RNA chimeric thereof, and SEQ ID NO:38, or an RNA equivalent or DNA/RNA chimeric thereof;
(iv) SEQ ID NO:39, or an RNA equivalent or DNA/RNA chimeric thereof, and SEQ ID NO:41, or an RNA equivalent or DNA/RNA chimeric thereof; and
(v) SEQ ID NO:42, or an RNA equivalent or DNA/RNA chimeric thereof, and SEQ ID NO:44, or an RNA equivalent or DNA/RNA chimeric thereof.
3 . The composition of claim 1 , wherein
the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (i) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:31, or an RNA equivalent or DNA/RNA chimeric thereof;
the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (ii) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:34, or an RNA equivalent or DNA/RNA chimeric thereof;
the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (iii) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:37, or an RNA equivalent or DNA/RNA chimeric thereof;
the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (iv) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:40, or an RNA equivalent or DNA/RNA chimeric thereof; or
the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (v) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:43, or an RNA equivalent or DNA/RNA chimeric thereof.
4 . The composition of claim 1 , wherein the Bpp-specific detection probe oligomer comprises the fluorescent label.
5 . The composition of claim 4 , wherein the Bpp-specific detection probe oligomer further comprises a non-fluorescent quencher.
6 . The composition of claim 3 , wherein the Bpp-specific detection probe oligomer comprises the fluorescent label.
7 . The composition of claim 6 , wherein the Bpp-specific detection probe oligomer further comprises a non-fluorescent quencher.
8 . The composition of claim 1 , further comprising a second amplification oligomer combination capable of amplifying a target region of a Bordetella pertussis (Bp) target nucleic acid.
9 . The composition of claim 3 , wherein the first and second Bpp-specific target-hybridizing sequences are target-hybridizing sequences of (i) and the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:31, or an RNA equivalent or DNA/RNA chimeric thereof.
10 . The composition of claim 1 , wherein the first and second Bpp-specific amplification oligomers comprise the first and second Bpp-specific target-hybridizing sequences of (a).
11 . The composition of claim 10 , wherein the first Bpp-specific target-hybridizing sequence is SEQ ID NO:30, or an RNA equivalent or DNA/RNA chimeric thereof.
12 . The composition of claim 11 , wherein the second Bpp-specific target-hybridizing sequence is SEQ ID NO:32, or an RNA equivalent or DNA/RNA chimeric thereof.
13 . The composition of claim 10 , wherein the Bpp-specific detection probe target-hybridizing sequence is SEQ ID NO:31, or an RNA equivalent or DNA/RNA chimeric thereof.
14 . The composition of claim 1 , wherein the first and second Bpp-specific amplification oligomers comprise the first and second Bpp-specific target-hybridizing sequences of (b).
15 . The composition of claim 1 , wherein the first and second Bpp-specific amplification oligomers comprise the first and second Bpp-specific target-hybridizing sequences of (c).
16 . A kit for determining the presence or absence of Bordetella parapertussis (Bpp) in a sample, said kit comprising the composition of claim 1 .
17 . A reaction mixture for determining the presence or absence of Bordetella parapertussis (Bpp) in a sample, said reaction mixture comprising the composition of claim 1 .
18 . The reaction mixture of claim 17 , further comprising at least one enzyme selected from the group consisting of a thermostable DNA polymerase, a reverse transcriptase, and an RNA polymerase.
19 . An aqueous formulation for determining the presence or absence of Bordetella parapertussis (Bpp) in a sample, said aqueous formulation comprising:
the composition of claim 1 , and
an organic buffer.
20 . A dried formulation for determining the presence or absence of Bordetella parapertussis (Bpp) in a sample, said dried formulation comprising:
the composition of claim 1 , and
a bulking agent.