The invention provides a plasmid shuttle vector characterized by being replicable in both of bacteria of the genus Brevibacillus and Escherichia coli, and by comprising a promoter capable of functioning in bacteria of the genus Brevibacillus and a DNA sequence(s) capable of functioning as a selective marker(s) for Escherichia coli and bacteria of the genus Brevibacillus, and a method of transforming bacteria of the genus Brevibacillus using the said plasmid shuttle vector. The plasmid shuttle vector enables the efficient transformation of bacteria of the genus Brevibacillus. Further, since plasmid shuttle vector pNCMO2 of the invention in particular has a gene expression control region which efficiently expresses and secretes an objective gene in bacteria of the genus Brevibacillus but significantly suppresses the expression in Escherichia coli, a recombinant gene expression plasmid can efficiently be constructed using Escherichia coli, and it is suited for protein production in bacteria of the genus Brevibacillus.