Expression in Bacillus subtilis of the gene for human urogastrone using synthetic ribosome binding sites

✍ Scribed by Flock, Jan-Ingmar ;Fotheringham, Ian ;Light, Janice ;Bell, Les ;Derbyshire, Roger

• Publisher: Springer
• Year: 1984
• Tongue: English
• Weight: 656 KB
• Volume: 195
• Category: Article
• ISSN: 0026-8925
• DOI: 10.1007/bf00332754

✦ Synopsis

A chemically synthesised gene coding for human urogastrone which was earlier cloned in E. coli (Smith et al. 1982) has now been cloned into expression vectors for Bacillus subtilis. Two types of constructs have been made, one giving production of methionyl-urogastrone and the other giving rise to a methionyl-urogastrone-beta galactosidase fusion polypeptide facilitating quantification of expression levels. The ribosome binding sites used in the expression plasmids are synthetically made oligonucleotides residing on short restriction fragments to allow easy replacement by other ribosome binding sites. Using "shuttle" vectors and constitutive promoters from Bacillus phages phi 105 and SPP1, we were able to detect levels of expression amounting to a few thousand molecules per cell during logarithmic growth in both E. coli and B. subtilis.