Detection of Influenza A Virus Neuraminidase and PB2 Gene Segments by One Step Reverse Transcription Polymerase Chain Reaction

We describe a single step reverse transcription polymerase chain reaction protocol that can be used to amplify part of the neuraminidase gene segment (segment 6) from all nine subtypes of influenza A virus. The method has also been applied to amplify gene segment 1 of influenza A, which encodes the basic polymerase protein 2 (PB2). The method combines the use of mixed base primers with a “touchdown” thermal cycling program and is applicable to a wide range of nucleic acid targets in which there is genetic variability in the regions complementary to the PCR primers.