Streptomyces tenebrarius H6 produces a complex of aminoglycoside antibiotics,such as apramycin,tobramycin and kanamycin B etc. To study the apramycin biosynthetic genes the genomic library from the Streptomyces tenebr...Streptomyces tenebrarius H6 produces a complex of aminoglycoside antibiotics,such as apramycin,tobramycin and kanamycin B etc. To study the apramycin biosynthetic genes the genomic library from the Streptomyces tenebrarius H6 was established using E.coli/Streptomyces shuttle vector pKC505 by in vitro packing.The probability of finding a specific gene from the library composed of 3 000 colonies was over 99 9%. According to the highly conserved sequence of the genes involved in 6\|deoxyhexose biosynthesis,primers were designed and 0 6kb fragment homologous to strE gene was obtained by PCR.30 positive clones were found from the genomic library of \%S.tenebrarius\% H6 with the 0 6kb fragment as a probe.Overlapped regions were localized by Southern hybridization and putative sugar related biosynthetic gene cluster was mapped by restriction enzyme digestions.An ORF of dTDP\|glucose\|4,6\|dehydratase gene consisted of 1,132bp,designated as apr E,was obtained and submitted to GenBank under the accession number of AF306787.A DNA sequence highly homologous to str L coding dTDP\|4\|dehydrorhamnose reductase was found linked with apr E gene.展开更多
文摘Streptomyces tenebrarius H6 produces a complex of aminoglycoside antibiotics,such as apramycin,tobramycin and kanamycin B etc. To study the apramycin biosynthetic genes the genomic library from the Streptomyces tenebrarius H6 was established using E.coli/Streptomyces shuttle vector pKC505 by in vitro packing.The probability of finding a specific gene from the library composed of 3 000 colonies was over 99 9%. According to the highly conserved sequence of the genes involved in 6\|deoxyhexose biosynthesis,primers were designed and 0 6kb fragment homologous to strE gene was obtained by PCR.30 positive clones were found from the genomic library of \%S.tenebrarius\% H6 with the 0 6kb fragment as a probe.Overlapped regions were localized by Southern hybridization and putative sugar related biosynthetic gene cluster was mapped by restriction enzyme digestions.An ORF of dTDP\|glucose\|4,6\|dehydratase gene consisted of 1,132bp,designated as apr E,was obtained and submitted to GenBank under the accession number of AF306787.A DNA sequence highly homologous to str L coding dTDP\|4\|dehydrorhamnose reductase was found linked with apr E gene.
