Granted Patent B1

US 9,598,739 · App. 14/288,691 · Granted Mar 21, 2017

Abstract

Claims (20)

1. A method for determining the presence of parvovirus B19 in a sample, the method comprising:

(a) performing an amplification step in which any parvovirus B19 nucleic acid present in the sample is amplified with at least one pair of primers comprising a first primer and a second primer, wherein the first primer comprises the nucleotide sequence as set forth in (SEQ ID NO:159); and

(b) performing a detection step which determines the presence or absence of an amplicon generated in step (a), comprising contacting the product of step (a) with a polynucleotide probe comprising the nucleotide sequence as set forth in (SEQ ID NO:160) or (SEQ ID NO:161), wherein positive detection of the amplicon indicates presence of a parvovirus B19 genotype 2 or 3 in the sample.

2. The method of claim 1 , wherein the second primer comprises the sequence as set forth in (SEQ ID NO:162).

3. The method of claim 1 , wherein the polynucleotide probe comprises a quencher molecule coupled to a pyrimidine ring of a thymine base.

4. The method of claim 3 , wherein the thymidine base is in nucleotide position 10 of (SEQ ID NO:160).

5. The method of claim 3 , wherein the quencher is a Black Hole Quencher®.

6. The method of claim 5 , wherein the Black Hole Quencher® is BHQ1.

7. The method of claim 1 , wherein the polynucleotide probe comprises a quencher molecule incorporated into the backbone of the probe.

8. The method of claim 7 , wherein the quencher replaces the nucleotide at position 10 of (SEQ ID NO:160).

9. A method for determining the presence of parvovirus B19 in a sample, the method comprising:

(a) performing an amplification step in which parvovirus B19 nucleic acid which may be present in the sample is amplified with at least one pair of primers comprising a first primer and a second primer, wherein the first primer comprises the nucleotide sequence as set forth in (SEQ ID NO:159) and the second primer comprises the nucleotide sequence as set forth in (SEQ ID NO:162); and

(b) performing a detection step which determines the presence or absence of an amplicon generated in step (a), comprising contacting the product of step (a) with a polynucleotide probe, wherein positive detection of the amplicon indicates presence of a parvovirus B19 genotype 2 or 3 in the sample.

10. The method of claim 9 , wherein the polynucleotide probe comprises the nucleotide sequence as set forth in (SEQ ID NO:160) or (SEQ ID NO:161).

11. The method of claim 9 , wherein the polynucleotide probe comprises a quencher molecule coupled to a pyrimidine ring of a thymine base.

12. The method of claim 11 , wherein the thymidine base is in nucleotide position 10 of (SEQ ID NO:160).

13. The method of claim 11 , wherein the quencher is a Black Hole Quencher®.

14. The method of claim 13 , wherein the Black Hole Quencher® is BHQ1.

15. The method of claim 9 , wherein the polynucleotide probe comprises a quencher molecule incorporated into the backbone of the probe.

16. The method of claim 15 , wherein the quencher replaces the nucleotide at position 10 of (SEQ ID NO:160).

Assignments (25)