Assay for
A region of the Trichomonas vaginalis AP65-1 gene has been identified which is useful for performing amplification assays to determine specifically whether T. vaginalis is present in the sample being tested. Oligonucleotides useful for performing thermal Strand Displacement Assay (tSDA) reactions on this gene are disclosed. The disclosed oligonucleotides can be used in an assay which is specific for multiple strains of T. vaginalis and which does not show cross reactivity with the genomes of other microorganisms or with human DNA.
1. A set of a plurality of primers and a probe for the detection of the AP65-1 gene of Trichomonas vaginalis comprising four primers and one probe wherein the four primers and one probe each have an oligonucleotide target binding sequence wherein the four oligonucleotide target binding sequences for the primers and the one oligonucleotide target binding sequence for the probe all hybridize within the SEQ ID NO:24 portion of the AP65-1 gene wherein the probe further comprises a detectable label attached to the probe oligonucleotide wherein the plurality of primers comprises a left bumper primer comprising SEQ ID NO: 7 and a right bumper primer comprising SEQ ID NO: a left primer comprising the target binding sequence consisting of SEQ ID NO: 9 and right primer comprising the target binding sequence consisting of SEQ ID NO: 10 and wherein the probe comprises a target binding sequence consisting of SEQ ID NO:11.
2. The probe set of claim 1 wherein the probe oligonucleotide sequence is SEQ ID NO:23.
3. The set of a plurality of primers and a probe for the detection of the AP65-1 gene of Trichomonas vaginalis according to claim 2 wherein the left primer is SEQ ID NO: 21 and the right primer is SEQ ID NO: 22.
4. The probe set of claim 1 wherein said detectable label is a fluorescence marker.
5. A kit for the amplification detection of Trichomonas vaginalis comprising:
a plurality of primers and a probe for the amplification detection of Trichomonas vaginalis that contains an AP65-1 gene (SEQ ID NO:1):
a) wherein the plurality of primers comprises a left bumper primer comprising SEQ ID NO: 7 and a right bumper primer comprising SEQ ID NO: 8, a left primer comprising the target binding sequence consisting of SEQ ID NO: 9 and right primer comprising the target binding sequence consisting of SEQ ID NO: 10 that bind within the highly conserved region identified by SEQ ID NO:24; and
b) wherein the probe in the set comprises a target binding sequence consisting of SEQ ID NO: 11 that binds within the highly conserved region identified by SEQ ID NO:24 and a detectable label attached to the probe oligonucleotide.
6. The kit of claim 5 further comprising reagents for performing a Strand Displacement Amplification (SDA) reaction using the set of primers and a probe.
7. The kit of claim 5 wherein the left primer is SEQ ID NO: 21 and the right primer is SEQ ID NO: 22 and the probe is SEQ ID NO: 23.