A bioluminescence assay to detect nitrification inhibitors released from plant roots: a case study with Brachiaria humidicola
A bioluminescence assay to detect nitrification inhibitors released from plant roots: a case study with Brachiaria humidicola
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DOI：
10.1007/s11104-006-9094-3
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发表时间：
2006-08
期刊：
影响因子：
4.9
通讯作者：
G. Subbarao;T. Ishikawa;Osamu Ito;K. Nakahara;Huoyan Wang;W. Berry
中科院分区：
文献类型：
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作者：
G. Subbarao;T. Ishikawa;Osamu Ito;K. Nakahara;Huoyan Wang;W. Berry
采用重组欧洲亚硝基单胞菌生物发光法检测和定量植物-土壤系统中天然硝化抑制剂。重组菌株n。在硝化过程中，由于从哈维弧菌引入的fluxabgenes的表达，欧洲豌豆产生了明显的双峰发光。该试验中产生的生物发光与NO2−产生高度相关（r2=0.94）。利用该试验，我们能够检测到由湿臂蒿（brachiaria humidicola(Rendle) Schweick）根产生的大量硝化抑制剂。我们建议将植物产生/释放的抑制活性称为“生物硝化抑制”（BNI），以区别于工业生产的抑制剂。根产生的BNI活性量以标准抑制剂烯基硫脲（AT）的作用为单位表示。在含有18.9 mM NH4+的实验中，0.22 μM AT的抑制作用被定义为一个AT活性单位。大量的BNI活性从b根释放出来。湿藓（15-25 AT单位g - 1根干wt day - 1）。BNI活性的释放与植株生长阶段和氮素含量有关。茎部氮水平与根系BNI活性释放呈显著正相关（r2=0.76）。抑制剂从mb中释放。湿根抑制土壤硝化作用。在55天的研究中，每克土壤添加20单位BNI完全抑制了NO3−的形成，并在50天内保持功能稳定。亚硝基单加氧酶和羟胺氧化还原酶的酶促途径均被mb释放的BNI活性有效阻断。humidicolaroots。该方法可用于植物根系BNI活性的表征和测定，为作物和牧场BNI性状的遗传开发提供了有力的工具。
A bioluminescence assay using recombinantNitrosomonas europaeawas adopted to detect and quantify natural nitrification inhibitors in plant–soil systems. The recombinant strain ofN. europaeaproduces a distinct two-peak luminescence due to the expression ofluxABgenes, introduced fromVibrio harveyi, during nitrification. The bioluminescence produced in this assay is highly correlated with NO2−production (r2=0.94). Using the assay, we were able to detect significant amounts of a nitrification inhibitor produced by the roots ofBrachiaria humidicola(Rendle) Schweick. We propose that the inhibitory activity produced/released from plants be termed ‘biological nitrification inhibition’ (BNI) to distinguish it from industrially produced inhibitors. The amount of BNI activity produced by roots was expressed in units defined in terms of the action of a standard inhibitor allylthiourea (AT). The inhibitory effect from 0.22 μM AT in an assay containing 18.9 mM of NH4+is defined as one AT unit of activity. A substantial amount of BNI activity was released from the roots ofB. humidicola(15–25 AT unit g−1root dry wt day−1). The BNI activity released was a function of the growth stage and N content of the plant. Shoot N levels were positively correlated with the release of BNI activity from roots (r2=0.76). The inhibitor/s released fromB. humidicolaroots suppressed soil nitrification. Additions of 20 units of BNI per gram of soil completely inhibited NO3−formation in a 55-day study and remained functionally stable in the soil for 50 days. Both the ammonia monooxygenase and the hydroxylaminooxidoreductase enzymatic pathways inNitrosomonaswere effectively blocked by the BNI activity released fromB. humidicolaroots. The proposed bioluminescence assay can be used to characterize and determine the BNI activity of plant roots, thus it could become a powerful tool in genetically exploiting the BNI trait in crops and pastures.