pHNC3, a suicide plasmid encoding luxAB and scaB genes in Tn5,can be transfered into Rhizobium through tri-parental mating and thenthe Rhizobium is labelled by luxAB. Since the levansucrasc activitycoded by sacB gene ...pHNC3, a suicide plasmid encoding luxAB and scaB genes in Tn5,can be transfered into Rhizobium through tri-parental mating and thenthe Rhizobium is labelled by luxAB. Since the levansucrasc activitycoded by sacB gene can cause the sucrose sensitivity on Gram-negativebacteria,the sacB gene allowed direct selection for the deletion or curingof plasmide, because only strains which no longer contain an activesacB gene are able to grow on media containing sucrose. This negativeselecting model can be used for direct labelling the mega-plasmid ofRhizobium,or even getting Rhizobium deriyatives partial deleted or curedtheir mega-plasmids.展开更多
The moblizable plashed pBF101 carrying Tn5-nifA of E. cloacae E26 was introduced into K. oxytoca NG13 with high frequency from donor E. coli S17-1 by mating. Tn5-nifA was transposed spontaneously from PBF101 into the ...The moblizable plashed pBF101 carrying Tn5-nifA of E. cloacae E26 was introduced into K. oxytoca NG13 with high frequency from donor E. coli S17-1 by mating. Tn5-nifA was transposed spontaneously from PBF101 into the genome of NG13 with a frequency of 6 ×10-5. Strain NG1394 was obtamed by selecting KmrCms colonies,which had lost plashed PBF101 and into which transposition of Tns-nifA had occurred. The integrahon of nifA gene into the genome in NG1394 was verified by agarose gel electrophoresis (Fig. 1) and curing of plasdrid with acridine orange. The synthesis of nitrogrnase in NG1394 was not repressed by (Table 1 ). Constitutive nifA could be stably maintained in NG1394 without antibiotic selection stress. borne nitrogenase activity was detected in NG1394 at 39℃ while NG13 (wild type) competely lost the activity at 39℃ (table 2).展开更多
文摘pHNC3, a suicide plasmid encoding luxAB and scaB genes in Tn5,can be transfered into Rhizobium through tri-parental mating and thenthe Rhizobium is labelled by luxAB. Since the levansucrasc activitycoded by sacB gene can cause the sucrose sensitivity on Gram-negativebacteria,the sacB gene allowed direct selection for the deletion or curingof plasmide, because only strains which no longer contain an activesacB gene are able to grow on media containing sucrose. This negativeselecting model can be used for direct labelling the mega-plasmid ofRhizobium,or even getting Rhizobium deriyatives partial deleted or curedtheir mega-plasmids.
文摘The moblizable plashed pBF101 carrying Tn5-nifA of E. cloacae E26 was introduced into K. oxytoca NG13 with high frequency from donor E. coli S17-1 by mating. Tn5-nifA was transposed spontaneously from PBF101 into the genome of NG13 with a frequency of 6 ×10-5. Strain NG1394 was obtamed by selecting KmrCms colonies,which had lost plashed PBF101 and into which transposition of Tns-nifA had occurred. The integrahon of nifA gene into the genome in NG1394 was verified by agarose gel electrophoresis (Fig. 1) and curing of plasdrid with acridine orange. The synthesis of nitrogrnase in NG1394 was not repressed by (Table 1 ). Constitutive nifA could be stably maintained in NG1394 without antibiotic selection stress. borne nitrogenase activity was detected in NG1394 at 39℃ while NG13 (wild type) competely lost the activity at 39℃ (table 2).
