COMPOSITIONS FOR USE IN IDENTIFICATION OF FRANCISELLA
First Claim
1. A composition, comprising at least one purified oligonucleotide primer pair that comprises forward and reverse primers, wherein said primer pair comprises nucleic acid sequences that are substantially complementary to nucleic acid sequences of two or more different bioagents belonging to the F. tularensis tularensis species, wherein said primer pair is configured to produce amplicons comprising different base compositions that correspond to said two or more different bioagents.
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Abstract
The present invention provides oligonucleotide primers, compositions, and kits containing the same for rapid identification of bacteria which are members of the Francisella genus by amplification of a segment of bacterial nucleic acid followed by molecular mass analysis.
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Citations
52 Claims
- 1. A composition, comprising at least one purified oligonucleotide primer pair that comprises forward and reverse primers, wherein said primer pair comprises nucleic acid sequences that are substantially complementary to nucleic acid sequences of two or more different bioagents belonging to the F. tularensis tularensis species, wherein said primer pair is configured to produce amplicons comprising different base compositions that correspond to said two or more different bioagents.
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20. A kit, comprising at least one purified oligonucleotide primer pair that comprises forward and reverse primers that are about 20 to 35 nucleobases in length, and wherein said forward primer comprises at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% sequence identity with a sequence selected from the group consisting of SEQ ID NOS:
- 1-2, 5-39, 75, 77, 79, and 81, and said reverse primer comprises at least 70% sequence identity with a sequence selected from the group consisting of SEQ ID NOS;
3-4, 40-74, 76, 78, 80, and 82.
- 1-2, 5-39, 75, 77, 79, and 81, and said reverse primer comprises at least 70% sequence identity with a sequence selected from the group consisting of SEQ ID NOS;
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21. A method of determining a presence of a Francisella in at least one sample, the method comprising:
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(a) amplifying one or more segments of at least one nucleic acid from said sample using at least one purified oligonucleotide primer pair that comprises forward and reverse primers that are about 20 to 35 nucleobases in length, and wherein said forward primer comprises at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% sequence identity with a sequence selected from the group consisting of SEQ ID NOS;
1-2, 5-39, 75, 77, 79, and 81, and said reverse primer comprises at least 70% sequence identity with a sequence selected from the group consisting of SEQ ID NOS;
3-4, 40-74, 76, 78, 80, and 82 to produce at least one amplification product; and(b) detecting said amplification product, thereby determining said presence of said Francisella in said sample. - View Dependent Claims (22, 23, 24, 25, 26)
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27. A method of identifying one or more Francisella bioagents in a sample, the method comprising:
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(a) amplifying two or more segments of a nucleic acid from said one or more Francisella bioagents in said sample with two or more oligonucleotide primer pairs to obtain two or more amplification products; (b) determining two or more molecular masses and/or base compositions of said two or more amplification products; and (c) comparing said two or more molecular masses and/or said base compositions of said two or more amplification products with known molecular masses and/or known base compositions of amplification products of known Francisella bioagents produced with said two or more primer pairs to identify said one or more Francisella bioagents in said sample. - View Dependent Claims (28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46)
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47. A system, comprising:
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(a) a mass spectrometer configured to detect one or more molecular masses of amplicons produced using at least one purified oligonucleotide primer pair that comprises forward and reverse primers, wherein said primer pair comprises nucleic acid sequences that are substantially complementary to nucleic acid sequences of two or more different Francisella bioagents; and (b) a controller operably connected to said mass spectrometer, said controller configured to correlate said molecular masses of said amplicons with one or more Francisella bioagent identities. - View Dependent Claims (48, 49, 50, 51, 52)
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Specification