Foot-and-mouth disease virus (FMDV) serotype Asia 1 is one of the most predominant endemic serotypes in China. Our previous study has generated a full-length cDNA clone (pBSAs) of an Asia 1 serotype FMDV (As1/CHA/05) isolated from bovine. To further study the properties of this virus, a mutant in the 3A region of the cDNA clone (pBSAs-3A10D), containing the deletion at position 93-102 of the 3A protein of As1/CHA/05, was generated by PCR and cloning. After synthesis of RNA in vitro and transfection, the recombinant rvAs-3A10D virus was recovered from BHK-21 cells. Characterization of the rvAs-3A10D revealed that the infectivity, immunoreactivity, and replication kinetics in BHK-21 and PK-15 cells and virulence in mice of the rvAs-3A10D were similar to that of its parent virus. Notably, while wild-type and recombinant viruses containing full-length sequence of the 3A replicated well in primary calf kidney cells, the mutant rvAs-3A10D failed to replicate in primary calf kidney cells in vitro. Apparently, the full-length sequence of 3A in As1/CHA/05 is a necessary component for its replication in calf kidney cells. The availability of this 3A deletion infectious cDNA clone may help in further investigating the virulent determinants of FMDV and potentially developing FMDV vaccines.