Optimized probes and primers and methods of using same for the binding, detection, differentiation, isolation and sequencing of influenza a; influenza b; 2009 influenza a/h1n1; and a 2009 influenza a/h1n1 rna sequence mutation associated with oseltamivir resistance

Abstract

Described herein are primers and probes useful for the binding, detecting, differentiating, isolating, and sequencing of influenza A, influenza B, 2009 influenza A/H1N1, and a 2009 influenza A/H1N1 RNA sequence mutation associated with oseltamivir resistance.

1 . A kit for detecting influenza A, and/or influenza B, and/or 2009 influenza A/H1N1, and/or oseltamivir-resistant 2009 influenza A/H1N1 and/or oseltamivir-sensitive 2009 influenza A/H1N1 in a sample, comprising one or more probes comprising a sequence selected from the group consisting of: SEQ ID NOS: 2 (influenza A); 5, 8, 15, 17, 20, 22, 25 (influenza B); 31, 37, 40, 43, 46 (2009 influenza A/H1N1); 56, 58, 63, 64 and 67 (oseltamivir-resistant 2009 influenza A/H1N1); 49, 54, 60, 65 and 66 (oseltamivir-sensitive 2009 influenza A/H1N1). 
     
     
         2 . The kit of  claim 1 , further comprising:
 a) at least one forward primer or primer pair comprising the sequence selected from the group consisting of: SEQ ID NOS: 1 (influenza A); 4, 7, 11, 13, 14, 19, 24, 27 (influenza B); 30, 33, 36, 39, 42, 45 (2009 influenza A/H1N1); 48, 51-53, 57, 61 (oseltamivir-resistant or sensitive 2009 influenza A/H1N1); and   b) at least one reverse primer or primer pair comprising the sequence selected from the group consisting of: SEQ ID NOS: 3 (influenza A); 6, 9, 10, 12, 16, 18, 21, 23, 26, 28, 29 (influenza B); 32, 34, 35, 38, 41, 44, 47 (2009 influenza A/H1N1); 50, 55, 59 and 62 (oseltamivir-resistant or sensitive 2009 influenza A/H1N1).   
     
     
         3 . The kit of  claim 2 , further comprising reagents for sequencing influenza A and/or influenza B and/or 2009 influenza A/H1N1 and/or oseltamivir-resistant 2009 influenza A/H1N1 and/or oseltamivir-sensitive 2009 influenza A/H1N1 in the sample. 
     
     
         4 . The kit of  claim 1 , further comprising a process control probe, process control forward primer and process control reverse primer selected from the group consisting of: Groups 30, 31 and 32 of Table 4. 
     
     
         5 . The kit of  claim 1 , wherein the one or more probes are labeled with different detectable labels. 
     
     
         6 . The kit of  claim 2 , wherein the at least one forward primer and the at least one reverse primer are selected from the group consisting of: Groups 1-29 of Table 4. 
     
     
         7 . A kit for detecting influenza A, and/or influenza B, and/or 2009 influenza A/H1N1, and/or oseltamivir-resistant 2009 influenza A/H1N1 and/or oseltamivir-sensitive 2009 influenza A/H1N1 and/or 2009 influenza A/H3N2 in a sample, comprising one or more probes comprising a sequence selected from the group consisting of: SEQ ID NOS: 2 (influenza A); 5, 8, 15, 17, 20, 22, 25 (influenza B); 31, 37, 40, 43, 46 (2009 influenza A/H1N1); 56, 58, 63, 64 and 67 (oseltamivir-resistant 2009 influenza A/H1N1); 49, 54, 60, 65 and 66 (oseltamivir-sensitive 2009 influenza A/H1N1); 101-123, 219-289, 363 and 364 (N2 of 2009 influenza A/H3N2). 
     
     
         8 . The kit of  claim 7 , further comprising:
 a) at least one forward primer or primer pair comprising the sequence selected from the group consisting of: SEQ ID NOS: 1 (influenza A); 4, 7, 11, 13, 14, 19, 24, 27 (influenza B); 30, 33, 36, 39, 42, 45 (2009 influenza A/H1N1); 48, 51-53, 57, 61 (oseltamivir-resistant or sensitive 2009 influenza A/H1N1); 77-100, 148-218, 361 and 362 (N2 of influenza A/H3N2); and   b) at least one reverse primer or primer pair comprising the sequence selected from the group consisting of: SEQ ID NOS: 3 (influenza A); 6, 9, 10, 12, 16, 18, 21, 23, 26, 28, 29 (influenza B); 32, 34, 35, 38, 41, 44, 47 (2009 influenza A/H1N1); 50, 55, 59 and 62 (oseltamivir-resistant or sensitive 2009 influenza A/H1N1); 124-147, 290-360, 365 and 366 (N2 of influenza A/H3N2).