Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (6): 1066-1073.doi: 10.3864/j.issn.0578-1752.2012.06.004

• TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY • Previous Articles     Next Articles

Isolation of Wheat Endophytic Diazotrophs and Determination of 1-Aminocyclopropane-1-Carboxylate Deaminase

 QIN  Bao-Jun, LUO  Qiong, GAO  Miao, HU  Hai-Yan, XU  Jing, ZHOU  Yi-Qing, SUN  Jian-Guang   

  1. 1.中国农业科学院农业资源与农业区划研究所/农业部作物营养与施肥重点实验室，北京 100081
  • Received:2011-03-29 Online:2012-03-15 Published:2011-12-29

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Abstract: 【Objective】 The objective of this study is to determine the ammount of wheat endophytic diazotrophs and screen for ACC (1-aminocyclopropane-1-carboxylate) deaminase activity from the diazotrophs, determine the phylogenetic and classific position of selected strains and prepare strains for microbial fertilizer production. 【Method】 Surface sterilization and nitrogen-free medium were used to isolate diazotroph and ACC was used as sole nitrogen source to screen strains with ACC deaminase activity. Nitrogenase activity was determined with acetylene reduction assay. 16S rDNA was amplified with PCR and analysed with MEGA software. Strain identification was carried out based on the morphology, physiology, biochemical test results and 16S rDNA analysis. 【Result】 The ammount of endophytic diazotrophs at jointing stage of wheat was (0.2-17.8)×105 cfu•g-1 fresh weight. Sixty endophytic diazotrophs with nitrogenase activity ranging 1-36 nmol C2H4/h•mg protein were isolated from wheat. Nine of the 60 endophytic diazotrophs were ACC deaminase positive, the range of enzyme activity is 0.87-9.32 µmol α-ketobutryric acid/h•mg protein. New isolate 9136 with nitrogenase activity 1.82 nmol C2H4/h•mg protein and ACC deaminase activity 9.32 µmol α-ketobutryric acid/h•mg protein was identified as Pseudomonas sp.. 【Conclusion】 About 105 cfu•g-1 (fresh weight) endophytic diazotrophs naturally colonized field grown wheat, some of these endophytic diazotrophs could produce ACC deaminase. A few strains showed relatively high ACC deaminase activity, and they might play a role in crop resistance to enviromental stress.

Key words: wheat, endophytic diazotrophs, ACC deaminase, PGPB

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