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Integration of Enterobacter cloacae E26 nifA gene into the genome of Klebsiella oxytoca NG13 by Tn5 transposition

Li Jie, Qiu Deyi, Shen Bingfu

Shanghai Institute of Plant Physiology,The Chinese Academy of Sciences. Shanghai 200032

Abstract

The mobilizable plasmid 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 obtained by selecting KmrCms colonies, which had lost plasmid pBF101 and into which transposition of Tn5-nifA had occurred.The integration of nifA gene into the genome in NG1394 was verified by agarose gel electrophoresis and curing of plasmid with acridine orange.The synthesis of nitrogenase in NG1394 was not repressed by NH~(+)_(-). Constitutive nifA could be stably maintained in NG1394 without antibiotic selection stress. Some nitrogenase activity was detected in NG1394 at 39℃ while NG13 (wild type) competely lost the activity at 39℃ .

Key words: Klebsiella oxytoca;nifA;Tn5;Transposition

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Citing This Article：

Li Jie, Qiu Deyi, Shen Bingfu. Integration of Enterobacter cloacae E26 nifA gene into the genome of Klebsiella oxytoca NG13 by Tn5 transposition. Acta Physiol Sin 1997; 49 (1): (in Chinese with English abstract).