Cloning Of Riboflavin Operon Of Bacillus Cereus And Expression In Bacillus Subtilis

In this work ¨two genomic fragments containing the complete riboflavin operonwere obtained from B.cereus ATCC 10987 and ATCC 14579 genomes bybioinformatics methods separately, which had good similarity with the DNA sequenceof riboflavin operon of B.subtilis. The products of the two riboflavin operon had atleast 98% similarity with the corresponding products of riboflavin operon of B.subtilisand B. amyloliquefaciens. The two fragments were amplified by PCR and cloned. It was found that both ofriboflavin operon were operative in E.coli. The riboflavin operon of B.cereus ATCC 14579 was chosen for further study.The evidences obtained from riboflavin auxotroph complementary experimentindicated that the riboflavin operon of B.cereus ATCC 14579 can express in B.subtilis.However, its expression in B.subtilis did not result in overproduction of riboflavin.The original promoter of the operon was replaced by the sacB promoter fromB.subtilis by a PCR method for its better expression. This substitution led to enhancedproduction of riboflavin under the fermentation condition used in this study. The yieldof riboflavin increased from 39.5mg/L to 61.7mg/L.