Bacillus pumilusBA06 has great potential for the production of alkaline proteases. To improve the protease yield, classical mutagenesis to combine the physical and chemical mutagens was performed to obtain a protease hyper-productive mutant SCU11. The full genome sequences of BA06 and SCU11 strains were assembled through DNA sequencing using the PacBio sequencing platform. By comparative genomics analysis, 147 SNPs and 15 InDels were found between these two genomes, which lead to alternation of coding sequence in 15 genes. Noticeable, the gene (kinA) encoding sporulation kinase A is interrupted by introducing a stop codon in its coding region in BA06. Interestedly, this gene is reversely corrected in SCU11. Furthermore, comparative transcriptome analysis revealed thatkinAand two positive regulatory genes (DegUandSpo0A) were upregulated in transcription in SCU11. In terms of the transcriptional data, upregulation of a phosphorylation cascade starting with KinA may enhance Spo0A phosphorylation, and thus activate expression of the geneaprE(encoding major extracellular protease) through repression of AbrB (a repressor ofaprE) and activation of SinI, an antagonist of SinR (a repressor ofaprE). In addition, the other genes involved in various metabolic pathways, especially of membrane transport and sporulation, were altered in transcription between these two strains. Conclusively, our transcriptome data suggested that upregulationdegUandspo0A,as well askinA,may at least partially contribute to the high production of alkaline protease in SCU11.