Abstract: To develop a simple and rapid method for detection of influenza A virus subtype H7N9，three pairs of specific primers were designed according to the conserved regions on the HA sequences of influenza A virus subtype H7，the NA sequences of influenza A virus subtype N9 and the M sequences of all influenza A viruses in GenBank，respectively. A triple polymerase chain reaction（ PCR） method was developed for detection of influenza A virus subtype H7N9 through optimization of triple PCR conditions，specificity test and sensitivity test. In this triple PCR，three specific fragments of the 304 bp of HA gene，the 160 bp of NA gene and the 458 bp of M gene could be amplified on the templates as influenza A virus subtype H7N9，but just one specific fragment of the 458 bp of M gene was amplifiedon the templates asthe other influenza A viruses，andthe triple PCR was negative for other viruses. Sensitivity test results showed that the detection limitof the triple PCR method was 0.6 pg of templates as H7N9 virus. Four hundred and seventy clinical samples were detected in the triple PCR，no influenza A virus subtype H7N9 was found. It was shown that the triple PCR method was rapid，sensitiveand specific，and it provided technical support for clinical test，effective prevention and control for influenza A virus subtype H7N9.