Detecting
There is described herein a method for specifically detecting Babesia species nucleic acid in a sample, which in one aspect comprises: (1) contacting a sample, said sample suspected of containing Babesia species nucleic acid, with at least two oligomers for amplifying a target region of a Babesia species target nucleic acid, wherein the at least two amplification oligomers comprise: (a) a first amplification oligomer comprising a first target-hybridizing sequence (i) that is from about 15 to about 33 contiguous nucleotides in length, is contained in the sequence of SEQ ID NO:66 and comprises SEQ ID NO:56 or 57; or (ii) that is from about 15 to about 33 contiguous nucleotides in length, is contained in the sequence of SEQ ID NO:96 and comprises SEQ ID NO:101; or (iii) that is from about 15 to about 33 contiguous nucleotides in length, is contained in the sequence of SEQ ID NO:97 and comprises SEQ ID NO:101; (iv) comprises or consists of SEQ ID NO:8; (v) comprises or consists of SEQ ID NO:83 and (b) a second amplification oligomer comprising a second target-hybridizing sequence that is from about 15 to about 33 contiguous nucleotides in length, and (i) is contained in SEQ ID NO:68 and comprises SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, or SEQ ID NO:85; or (ii) is contained in SEQ ID NO:67 and comprises SEQ ID NO:45 or SEQ ID NO:52; or (iii) is contained in SEQ ID NO:70 and comprises SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, or SEQ ID NO:51; (2) performing an in vitro nucleic acid amplification reaction, wherein any Babesia target nucleic acid present in said sample is used as a template for generating an amplification product; and (3) detecting the presence or absence of the amplification product, thereby indicating the presence or absence of Babesia species target nucleic acid in said sample.
1. A combination of at least two oligomers for determining the presence or absence of Babesia in a sample, said oligomer combination comprising first and second amplification oligomers for amplifying a target region of Babesia target nucleic acid, wherein
(a) the first amplification oligomer comprises a first target-hybridizing sequence that consists of SEQ ID NO:8, wherein the first amplification oligomer is a promoter primer or promoter provider further comprising a promoter sequence located 5′ to the first target-hybridizing sequence; and
(b) the second amplification oligomer comprises a second target-hybridizing sequence that consists of SEQ ID NO:34.
2. The combination of claim 1 , wherein the promoter sequence is a T7 promoter sequence.
3. The combination of claim 2 , wherein the first amplification oligomer nucleotide sequence consists of SEQ ID NO:7.
4. The combination of claim 1 , further comprising at least one capture probe oligomer, wherein the at least one capture probe oligomer comprises a target-hybridizing sequence covalently attached to a sequence or moiety that binds to an immobilized probe, wherein said target-hybridizing sequence consists of SEQ ID NO:44, SEQ ID NO:88, or SEQ ID NO:90.
5. The combination of claim 4 , wherein the capture probe oligomer sequence comprises or consists of SEQ ID NO:43, SEQ ID NO:87, or SEQ ID NO:89.
6. The combination of claim 1 , further comprising a detection probe oligomer configured to specifically hybridize to a Babesia amplification product amplifiable by the first and second amplification oligomers.
7. The combination of claim 6 , wherein the detection probe oligomer comprises a detection probe target-hybridizing sequence consisting of the sequence selected from the group consisting of: SEQ ID NO:37, SEQ ID NO:42, SEQ ID NO:91, SEQ ID NO: 92, SEQ ID NO:93, and combinations thereof.
8. The combination of claim 7 , wherein the detection probe comprises a fluorescent label and a quencher.
9. The combination of claim 1 , wherein the at least two amplification oligomers further comprise:
(a) a third amplification oligomer comprising a third target-hybridizing sequence that consists of SEQ ID NO:83;
(b) a fourth amplification oligomer comprising a fourth target-hybridizing sequence that consists of SEQ ID NO:84; and,
(c) a fifth amplification oligomer comprising a fifth target-hybridizing sequence that consists of SEQ ID NO:86.
10. The combination of claim 9 , further comprising at least one capture probe oligomer, wherein the at least one capture probe oligomer comprises a target-hybridizing sequence covalently attached to a sequence or moiety that binds to an immobilized probe, wherein said target-hybridizing sequence consists of SEQ ID NO:44, SEQ ID NO:88, or SEQ ID NO:90.
11. The combination of claim 1 , further comprising a detection probe oligomer, wherein the detection probe oligomer comprises a detection probe target-hybridizing sequence consisting of a sequence selected from the group consisting of SEQ ID NO: 91, SEQ ID NO:92, and SEQ ID NO:93.
12. A combination of at least two oligomers for determining the presence or absence of Babesia in a sample, said oligomer combination comprising first and second amplification oligomers for amplifying a target region of Babesia target nucleic acid, wherein
(a) the first amplification oligomer comprises a first target-hybridizing sequence that consists of SEQ ID NO:8, wherein the first amplification oligomer is a promoter primer or promoter provider further comprising a promoter sequence located 5′ to the first target-hybridizing sequence;
(b) the second amplification oligomer comprises a second target-hybridizing sequence that consists of SEQ ID NO:34; and
(c) a detection probe oligomer configured to specifically hybridize to a Babesia amplification product amplifiable by the first and second amplification oligomers, wherein the detection probe oligomer comprises a 2′ methoxy modification on at least one nucleotide residue member of the nucleotide sequence.
13. A combination of at least two oligomers for determining the presence or absence of Babesia in a sample, said oligomer combination comprising first and second amplification oligomers for amplifying a target region of Babesia target nucleic acid, wherein
(a) the first amplification oligomer comprises a first target-hybridizing sequence that consists of SEQ ID NO:8, wherein the first amplification oligomer is a promoter primer or promoter provider further comprising a promoter sequence located 5′ to the first target-hybridizing sequence;
(b) the second amplification oligomer comprises a second target-hybridizing sequence that consists of SEQ ID NO:34; and
(c) a detection probe oligomer configured to specifically hybridize to a Babesia amplification product amplifiable by the first and second amplification oligomers, wherein the detection probe comprises a chemiluminescent label or a fluorescent label.