DETECTION OF TN5-LIKE SEQUENCES IN KANAMYCIN-RESISTANT STREAM BACTERIA AND ENVIRONMENTAL DNA

被引:31
作者
LEFF, LG [1 ]
DANA, JR [1 ]
MCARTHUR, JV [1 ]
SHIMKETS, LJ [1 ]
机构
[1] SAVANNAH RIVER ECOL LAB, AIKEN, SC 29802 USA
关键词
D O I
10.1128/AEM.59.2.417-421.1993
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
Resistance to kanamycin and neomycin in the bacterial assemblage of a coastal plain stream was detected by growth of colonies on media containing antibiotics. Three of 184 kanamycin-resistant colonies hybridized with a probe containing the nptII gene from transposon Tn5; the nptII gene encodes the enzyme neomycin phosphotransferase and conveys resistance to kanamycin and neomycin. In one of these isolates, the homologous gene was cloned and shown to confer resistance to a kanamycin-sensitive Escherichia coli strain. Since enumeration of bacteria by acridine orange direct counts revealed that less than 0.2% of the bacteria present were cultivated, direct examination of environmental DNA was used to assess abundance of sequences that hybridize to the nptII gene. To examine the resistance potential of bacteria that were not cultured, total DNA was extracted from environmental samples and hybridized with specific probes. The relative amount of eubacterial DNA in each sample was determined by using a eubacterial specific rDNA probe. Then, the abundance of sequences that hybridize to the eubacterial neomycin phosphotransferase gene was determined by hybridization and expressed relative to the total eubacterial DNA in the assemblage. Relative gene abundance was significantly different among assemblages from different habitats (leaves, midchannel sediments, and bank sediments) but did not differ among stream sites.
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页码:417 / 421
页数:5
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