In 2010, an H4N6 avian influenza virus (AIV) was isolated and identified from healthy ducks in a waterfowl market in Shandong Province in the northern China. This virus was named A/duck/Shandong/1/2010 (H4N6) (DK/SD/1/2010 hereafter). The gene sequence of the virus was determined, and genetic and evolutionary analyses were conducted by combining related sequences in GenBank. Results indicated that seven genes of DK/SD/1/2010 (PB2, PB1, PA, HA, NP, M, and NS) originate from the Eurasian lineage. Another gene, the NA gene, originated from both the Eurasian and the North American lineages. The amino acid sequence near the cleavage site of DK/SD/1/2010 HA (PKKASR↓GLF) corresponded to the characteristics of AIV of low pathogenicity. Animal inoculation tests showed that the virus cannot replicate in chickens and mice. Therefore, DK/SD/1/2010 may be a recombinant virus formed by influenza virus genes from different sources through complicated restructuring and evolution in ducks that is avirulent to chickens and mice.