Abstract: Based on the mechanism of ligase-dependent RT-PCR assay, we established RT-PCR assay to detect avian tembusu virus which still prevails in most poultry of China.The results showed that it would specifically detect tembusu viruses isolated from different avian, such as laying duck, meat duck, goose and hens.The sensitivity of this assay is equivalent to that of conventional RT-PCR.To test the reliability of ligase-dependent RT-PCR assay, six tissue samples collected from experimental animals and 140clinical samples collected from Fujian, Jiangxi, Zhejiang, and Guangdong province during 2011-2012were detected.All of the six tissue samples were positive, while only 20of the 140clinical samples were tested to be positive by this method.The above results indicated that the ligase-dependent RT-PCR assay was a sensitive, specific and rapid diagnosis method for detecting avian Tembusu virus.