Abstract: Taking Corynebacterium glutamicum MD106 as a starting strain applied to produce Lleucine, the mutant strain MD106ΔpanBC/ΔalaT from the double genenegative of panBC and alaT was structured by using the methods of overlap extension PCR and suicide vectormediated homologous recombination, and the fermentation index was measured through the ermentation test of the starting strain and the recombinant strain. The results showed that the Lleucine yield of the recombinant strain was 7.91g/L by Amino acid analyzer after40 h fermentation, and it increasing by 43.3% comparing with that of the original strain MD106, and mainly miscellaneous acid——alanine reduced by more than 80%, the total ration of miscellaneous acid decreased 55.6% compared with the original strain.