To construct a promoter probe vector, pBE-bgaB, to screen strong promoters from Bacillus amyloliquefaciens. 266 colonies containing active promoter elements from the genomic DNA of B. amyloliquefaciens were identified. Among these, promoter P41 exhibited the strongest beta-Gal activity in Escherichia coli and B. amyloliquefaciens. Sequence analysis showed that promoter P41 contained P (ykuN) , a ykuN gene encoding flavodoxin. Optimization of the ribosome-binding site from P41 to P-382 improved beta-Gal activity by similar to 200%. A new strong promoter for protein expression and genetic engineering of Bacillus species.