Abstract: Total RNA was extracted from tissue samples of FMDV China99/S strain and four overlapping fragments (A， B， C and E) were amplified. The pP_ⅠcDNA3.1（+）vector with RNA polymerase Ⅰand A， B， C， E fragments were digested respectively by restriction endonucleases(A(BsmBⅠ/XmaⅠ)，B(XmaⅠ/ BamHⅠ)， C(HpaⅠ/KpnⅠ)，E(BamHⅠ/NotⅠ))， and then ligated to produce recombinant plasmid of pP_ⅠFMDVcDNA3.1（+）. The analysis of nucleotide sequences showed that the genome of swine FMDV China99/S strain is 8 116 nucleotides in length(including Poly(C) and Poly(A) tracts)， and the open reading frame (ORF， 6 318 nucleotides which encode 2 106 amino acids) is flanked by a 5′non-coding region (5′NCR) of 1 064 nucleotides (including 41 C residues Poly(C) tract) and a 3′NCR of 131 nucleotides (including the Poly(A) tail at least 38 A residues). Recombinant plasmid of pP_ⅠFMDVcDNA3.1（+） was transferred into BHK-21 cells with liposome and typical FMDV pathological changes were observed in subculture. The electron microscope,CPE,animal test,RT-PCR and sequencing analysis verified that FMDV China99/S was rescued.