Table 1. Primer sequences designed for NA subtyping and the expected PCR products.
| NA subtype | Primer sequence | Product length (bp) + | TM value ++ |
|---|---|---|---|
| N1-foward | 5'-TAGACTGCATGAGGCCTTGCTTCTG-3' | 137 | 78.2~79.2 |
| N1-reverse | 5'-CACCGTCTGGCCAAGACCAACCTAC-3' | ||
| N2-foward | 5'-ATGTTATCAATTTGCACTTGGGCAG-3' | 149 | 77.1-77.8 |
| N2-reverse | 5'-CATGCTATGCACACTTGTTTGGTTC-3' | ||
| N3-foward | 5'-ATGATGTCTCTTGGACAAGCAATAG-3' | 104 | 74.8-76.2 |
| N3-reverse | 5'-TGGGCATAAACCCAATGTTGGAACC-3' | ||
| N4-foward | 5'-AAATCATAACCATCGGTAGTGCGAG-3' | 194 | 76.8-78.6 |
| N4-reverse | 5'-TATAGTTGTTCTGCACATTGGTGAC-3' | ||
| N5-foward | 5'-CATTTGTGGCATGTGGTCCCACGGA-3' | 147 | 76.6-77.2 |
| N5-reverse | 5'-AGGCATTGGGTGAAGATCCTAATGG-3' | ||
| N6-foward | 5'-GCAAATAGACCAGTAATCACTAT-3' | 153 | 77.9-78.9 |
| N6-reverse | 5'-CCAGGATCTGGGTTTCCTCCTGTTA-3' | ||
| N7-foward | 5'-AGCCAAGTATGTTTGGTGGACGAGC-3' | 111 | 79.2-80.3 |
| N7-reverse | 5'-TTACGAAAAGTATTGGATTTGTGCC-3' | ||
| N8-foward | 5'-TAATGAGTGTAGAAATAGGGCAATC-3' | 127 | 78.9-79.6 |
| N8-reverse | 5'-GGAATCAGGGCCCGTTACTCCAA-3' | ||
| N9-foward | 5'-ATCGTATTAAACACTGACTGGAGTG-3' | 171 | 78.2-78.9 |
| N9-reverse | 5'-ATTCTGTGCTGGAACACATTGATAC-3' |
+
The expected length of PCR product was calculated according to reference sequences of corresponding NA subtype in NCBI influenza database.
++
The range of TM value was analyzed by serially diluted DNA or RNA template of different NA subtype in Real-time PCR or Real-time RT-PCR.