低氮胁迫下苦荞根际土壤纤维素酶活性的响应机制：荧光光谱法测定

土壤纤维素酶参与土壤中养分的循环过程，分解纤维素为小分子糖分微生物生命活动提供能源物质，并且对生态系统有重要的指示作用。如何快速、精确的测定纤维素酶活性对土壤生态系统诊断、恢复等有重要指示意义。自然界中不同的氮肥处理及种植不同的苦荞品种均会对土壤纤维素酶活性产生影响。利用德国进口全自动进样酶标仪(InfiniteR 200 PRO)，以4-Methylumbelliferyl β-D-cellobiopyranoside (Cel) 纤维素荧光物质为底物，将不同耐瘠性苦荞(迪庆苦荞-耐低氮；黑丰1号-不耐低氮)根际土壤以悬液形式用排枪吸取微量和荧光底物混合加入到96孔微孔板进行充分反映、培养测定土壤中纤维素酶活性，其中将对照板和样品放在同一个板块上，减少培养空间占用和操作时间。结果表明，除了苦荞成熟期的常氮处理，苗期和开花期的低氮处理条件下，纤维素酶活性均表现为迪庆苦荞根际土壤显著高于黑丰1号，迪庆根际纤维素酶活性在苗期低氮处理下高于黑丰39.50%；纤维素酶活性在迪庆苦荞的开花期和成熟期氮处理间有显著性差异，表现为常氮高于其他处理，而黑丰1号仅在开花期时常氮处理和其他处理之间有显著差异；灭菌处理酶活性显著低于常氮处理的54.29%，说明迪庆苦荞根际纤维素酶一部分来源于微生物的贡献，养分充足时微生物的活性更高，贡献越大。荧光光谱法分析出的纤维素酶活性表明耐瘠性品种迪庆苦荞可以通过增加根际土壤酶活性的方法来抵抗外界环境的胁迫，面对黄土高原养分贫瘠的土壤，可以考虑选种耐贫瘠的品种来降低成本增加收益。试验过程中，荧光标记的酶标仪测定方法与传统的测定方法相比更加快捷、准确和节约，同样适合大批量样品测定，可以为未来精准农业生产，测土配方施肥提供及时的数据支持。

Fluorescence Spectrometric Determination of Cellulase in the rhizosphere of Buckwheat Soil under Low Nitrogen Stress

Cellulasel plays an very important role in the ecosystem and the nutrient cycling. It can decomposite cellulose to small molecules of sugar and provide energy for biological activities. How to measure cellulase activity quickly and accurately is of great significance for the diagnosis and restoration for soil ecosystem. Different nitrogen treatments and cultivate would influenced enzyme activity. This study use automatic TECAN Infinite 200 Multi-Mode Microplate Reader (made in Germany) bombined with 96 microplates and fluorescence (4-Methylumbelliferyl β-D-cellobiopyranoside (Cel)) detection method to detect the activity of cellulasel under different tartary buckwheat (DiQing-low nitrogen toleration, HeiFeng 1-low nitrogen intoleration). During the process, the experiment use less substrate and put the control and sample together in the same plat,then reduced the culture space and operation time. The results showed that except the normal nitrogen treatment under the maturation stage, cellulasel was higher in DiQing than in HeiFeng 1 in the stage of seeding and flowering. In addition, the activity of cellulase in root trace of Diqing was higher than that of Heifeng 39.50% under low nitrogen treatment at seedling stage. Cellulasel had a significant difference among nitrogen treatments in the flowering and maturation stage, much higher activity in normal treatment when compared with the others under DiQing, but Heifeng only showed the same trend in flowering stage. The enzymatic activity of sterilizing treatment was significantly lower than that of normal nitrogen treatment of 54.29%, which indicated that some of cellulase in root trace of Diqing Tartary buckwheat originated from the contribution of microorganism, and the activity of microorganism was higher when the nutrient was sufficient. The analysis of cellulase activity by fluorescence spectrum showed that the resistant variety Diqing Tartary buckwheat could resist the stress of external environment by increasing the activity of soil enzyme in root trace. While in the poor nutrient soil in Loess Plateau, maybe selecting barren-tolerant varieties would reduce costs and increase the benefits for farmers. Compared with traditional method, fluorescence microplate fluorescence microplate method is quick, accurate and economical. It is also suitable for mass sample determination, which can provide timely data support for foreign precision agriculture production and soil testing formula fertilization in the further.