Cloning of the rfb gene region of Shigella dysenteriae 1 and construction of an rfb-rfp gene cassette for the development of lipopolysaccharide-based live anti-dysentery vaccines.
Recent studies have shown that determinants for the production of O antigen lipopolysaccharide in Shigella dysenteriae 1 are distributed over two distinct genetic elements, the chromosome and a 9 kb plasmid designated pHW400. In this communication, we describe the cloning of all determinants necessary for S. dysenteriae 1 O antigen production in E. coli K-12 and their combination in a single plasmid. An RP4::miniMu R-prime plasmid, R-prime 40, containing the his-rfb (histidine biosynthesis-lipopolysaccharide biosynthesis) gene region of the Shigella dysenteriae 1 chromosome was generated. E. coli K-12 bacteria containing R-prime 40 and pSS8, a transposon Tn5-tagged derivative of pHW400, produced lipopolysaccharide indistinguishable from that of S. dysenteriae 1. Small DNA fragments containing the rfb gene cluster and the rfp gene were subcloned from R-prime 40 and pSS8 and subsequently combined in vector pACYC184 to produce pSS37. This latter plasmid when introduced by transformation into E. coli K-12 provoked the formation of S. dysenteriae 1 O-specific lipopolysaccharide, a feature that suggests it may be useful in the construction of LPS-based live vaccines against the Shiga bacillus.
Sturm
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S
Department of Medical Biochemistry, University Medical Centre, University of Geneva, Switzerland.
Timmis
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KN
D000942:Antigens, Bacterial / Q000235:genetics; D001428:Bacterial Vaccines / Q000302:isolation & purification; D003001:Cloning, Molecular; D004405:Dysentery, Bacillary / Q000517:prevention & control; D005798:Genes, Bacterial; D006801:Humans; D008070:Lipopolysaccharides / Q000235:genetics / Q000276:immunology; D019081:O Antigens; D010957:Plasmids; D012762:Shigella dysenteriae / Q000235:genetics* / Q000276:immunology
D000942:Antigens, Bacterial; D001428:Bacterial Vaccines; D008070:Lipopolysaccharides; D019081:O Antigens