Detection and analysis of influenza virus

• US 7,972,786 B2
• Filed: 07/06/2007
• Issued: 07/05/2011
• Est. Priority Date: 07/07/2006
• Status: Expired due to Fees

• Current Assignee: Brandeis University
• Original Assignee: Brandeis University
• Inventors: Hartshorn, Cristina, Rice, John E., Pierce, Kenneth E., Reis, Arthur Henry Jr., Sanchez, J. Aquiles, Wangh, Lawrence J.
• Primary Examiner(s): Horlick; Kenneth R.
• Assistant Examiner(s): Tung; Joyce
• Application Number:
  US11/822,536
• Publication Number:
  US 20090081648A1
• Time in Patent Office: 1,460 Days
• Field of Search: 536/23.1, 536/24.32, 435/6
• US Class Current: 435/6.12
• CPC Class Codes: C12Q 1/701 Specific hybridization probes

Abstract

An assay comprising more than one primer pair and more than one detection probe, a low copy number synthetic amplicon corresponding to each of the primer pairs. The assay can detect and distinguish between various sub-types and strains of an influenza virus using any suitable nucleic acid amplification technique. Related kits and methods are also described.

Claims

1. A kit for performing an assay comprising more than one primer pair, more than one detection probe, and a low copy number synthetic amplicon corresponding to each of the primer pairs, wherein the primer pairs are selected from the group consisting of:

a) CAGCACCGTCTGGCCAAGAC (SEQ ID NO;

     5), and GCAATAACTGATTGGTCAGG (SEQ ID NO;

     6);

b) CGTTGTATGACCAGAGATCTATTTTAGTGTCCT (SEQ ID NO;

     7), and CCATCAGATTGAAAAAGAATTCT (SEQ ID NO;

     8);

c) CAGGAGGTCTATATTTGGTTCCATTGGC (SEQ ID NO;

     9), and CGGTGGATTAAACAAAAGCA (SEQ ID NO;

     10);

d) CCCAATACAGGGGACATCACATTTCTTG (SEQ ID NO;

     11), and CATGGGCTGACAGTGAT (SEQ ID NO;

     12);

e) GGTGACAGGATTGGTCTTGTCTTTAGC (SEQ ID NO;

     13), and CTAACCGAGGTCGAAAC (SEQ ID NO;

     14)

f) GATGCAGCTTTTGCCTTCAACAGAG (SEQ ID NO;

     15), and GGTCCAACCCTAATTCCAA (SEQ ID NO;

     16); and

g) CCTCCCTCTATAAAACCTGCTATAGCTCCAAA (SEQ ID NO;

     17), and CGACTGGGCTCAGAAA (SEQ ID NO;

     18);

wherein the detection probe is a molecular beacon probe selected from the group consisting of;

a) Texas Red-CGCGACTAGGGAACTCGCTCGCG (SEQ ID NO;

     19)-Dabsyl,

b) CY3-CGCGGATTGGCTTTTTACTTTCTCACCGCG (SEQ ID NO;

     20)-Dabsyl,

c) FAM- GGCGGATGCTGCTCCCACTACCGCC (SEQ ID NO;

     21)-Dabsyl,

d) CY5-CGCTGAAAGCGTTTCTCGAGGTCCTG (SEQ ID NO;

     22)-BHQ1,

e) Texas Red-GCGAGTTTGCATGTTCTCCTGTCTCGC (SEQ ID NO;

     23)-Dabsyl,

f) CY5-GCGAGTTTGCATGTTCTCCTGTCTCGC (SEQ ID NO;

     23)-Dabsyl,

g) CY3-GCGCTATAGAGAGAACAGCGC (SEQ ID NO;

     25)-Dabsyl, Tm=33.8

h) FAM-GGCCGCCTATTACCTCTCGGCC (SEQ ID NO;

     26)-Dabsyl, and

i) CY5-CGCTGAAAGCGTTTCTCGAGGTCCTG (SEQ ID NO;

     22)-BHQ1.

2. The kit according to claim 1

wherein the primer pairs additionally include at least one pair selected from the group consisting of:

h) GGATAGACCAGCTACCATGATTGCC (SEQ ID NO;

     1), and GTGGAGTAAAATTGGAATCAATAGG (SEQ ID NO;

     2); and

i) CACCCGTTTCCTATTTCTTTGGCATTATTC (SEQ ID NO;

     3), and CCATGACTCCAATGTGAAG (SEQ ID NO;

     4);

wherein the detection probe additionally includes at least one molecular beacon probe selected from the group consisting of;

j) Texas Red-CGCGACTAGGGAACTCGCTCGCG (SEQ ID NO;

     19)-Dabsyl, and

k ) CY3-CGCGGATTGGCTTTTTACTTTCTCACCGCG (SEQ ID NO;

     20)-Dabsyl.

3. The kit of claim 1, wherein the low copy number synthetic amplicon comprises approximately twenty copies of the synthetic amplicons per primer pair.

4. The kit of claim 2, comprising all of primer pairs a)-i).

5. The kit of claim 1, further comprising reagents for an amplification reaction.