The invention relates to the technical field of biological engineering, and especially relates to a ketogulonigenium vulgare engineering strain, a preparation method and an application thereof. The ketogulonigenium vulgare engineering strain comprises a key gene during an amino acid synthesis process; and the key gene in the amino acid synthesis process is in the key gene during a threonine synthesis process, the key gene during a proline synthesis process and the key gene during a histidine synthesis process. By constructing the deleted gene during the amino acid synthesis process in the bacterial strain, when individual fermentation of the ketogulonigenium vulgare engineering strain is carried out, sorbose can better conversed to ketogulonigenium vulgare; when ketogulonigenium vulgare engineering strain and the endophytic bacillus are subjected to mixed fermentation, the fermentation period is reduced, compared with the original bacterial strain, the period is shortened by about 10 h, the time cost is saved; from the yield aspect, the yield is increased by about 20% by comparing with the original strain; effect of companion fungus dependence is reduced when mixed fermentation is carried out, amount of the companion fungus which is Bacillus spp is reduced, and the cost and pollution are reduced.