Abstract: The specification describes a process for producing human proinsulin in Escherichia coli (E. coli) using gene manipulation technology. The process can provide for human proinsulin in high yields by a novel expression vector having strong regulatory elements of an insulin gene and a stable recombinant gene product. The expression vector of the present invention is characterized in that: 1) it has an 11 amino acid leader peptide containing six threonines in order to ensure an intracellular stability of proinsulin fusion protein, 2) it contains two copies of a DNA expression cassette each comprising a strong lambda P.sub.R promoter, a lac ribosome binding site, a proinsulin gene with a 17 amino acid leader peptide sequence containing a DNA sequence encoding (Thr).sub.6, and a strong fd phage transcription terminator (combination of phage fd terminator and translation stop codon), etc.