Based on the reported gene sequences, the segments containing 2-keto aldose reductase (2-KRA and B) genes were amplified by PCR from the plasmids and Erwinia sp. SCB125 each for gene expression and gene knocking out. Then cloning them into expression vector pBL and successfully expressing them with high enzyme activity in E. coli DH5 alpha. After their enzyme activities were proved, the work on gene knock out followed. Introducing the knock-out vector which distribute unstably during the cell division to the host strains Erwinia sp. SCB125. Screening firstly by the positive marker, one resistance which resulted from the expression of the resistance gene inserting inside the reductase genes and the negative marker, another resistance which outside the reductase genes in the vector. The strains selected out will be tested by further study. This work was the bases of blocking the pathway metabolism and constructing a recombinant strain that can produce 2-KLG directly from D-glucose by one-step fermentation.