植物根尖分生组织传感器的构建及其对尿素传感动力学研究

Construction of plant root tip mesenchymal sensors and their effects on urea sensing dynamics

为探寻植物根尖组织对尿素的传感能力和两者相互作用的动力学规律。以海藻酸钠—淀粉凝胶作固定剂,将玉米、辣椒、花椰菜和黄瓜的根尖分生组织固定到2片核微孔膜之间,制成“三明治”式尿素传感膜,然后将其固定并密封到玻碳电极上,制成植物根尖分生组织传感器。通过电化学工作站和时间—电流曲线法测定尿素与玉米、辣椒、花椰菜和黄瓜的根分生组织传感器互作所产生电化学信号的变化。与对照组相比,4种植物根尖分生组织传感器对不同浓度的尿素均呈现明显的函数关系。根分生组织传感器对不同浓度的尿素传感动力学测定结果表明:玉米、辣椒、花椰菜和黄瓜分别在10^-8~10^-4、10^-16~10^-6、10^-19~10^-10和10^-20~10^-10 mol·L^-1的尿素范围内呈现类似于酶和底物响应的酶促反应特征(联动变构效应)。进一步分析显示:玉米、辣椒、花椰菜、黄瓜和尿素的联动变构常数Ka(类似于酶—底催化动力学的米氏常数Km)分别为:7.1970×10^-9、4.5370×10^-16、9.9085×10^-20和6.4628×10^-21 mol·L^-1,表明玉米对尿素的传感能力至少比其他3种植物差7个数量级以上。在以尿素为唯一外源氮营养的条件下培养玉米和辣椒,实验结果证明,在低于玉米传感尿素能力下限(1×10-10 mol·L^-1)时,玉米10~15 d枯黄死亡,而辣椒可以正常生长。证明联动变构常数Ka反映了植物根尖分生组织对尿素的真实传感能力,而且玉米对尿素的传感存在某种程度上的缺陷。

The plant root tip tissue sensor was constructed to quantitatively measure the root tip tissue's ability to sense urea and to explore the dynamics of the interaction between the two.In this study,sodium alginate-starch gel was used as a fixative.The meristems of corn,pepper,cauliflower and cucumber were fixed between two nuclear microporous membranes to make a“sandwich”urea sensing membrane.It was fixed and sealed on a glassy carbon electrode to make a plant root tip meristem sensor.Electrochemical workstations and time-current curve methods were used to determine the changes in electrochemical signals produced by the interaction of urea with root meristem sensors of corn,pepper,cauliflower,and cucumber.Compared with the blank control group,the root-tip meristem sensors of four plants showed a clear functional relationship to different concentrations of urea.Corn,pepper,cauliflowerand cucumber exhibited enzymatic response characteristics(linked allosteric effect)was similar to enzyme and substrate responses in the range of 10^-8-10^-4,10^-16-10^-6,10^-19-10^-10 and 10^-20-10^-10mol·L^-1 urea.Further analysis showed that the linked allosteric constants Ka(similar to the Mie constants Km of enzyme-bottom catalytic kinetics)of corn,pepper,cauliflower,cucumberand urea were:7.1970×10^-9,4.5370×10^-16,9.9085×10^-20 and 6.4628×10^-21mol·L^-1.It was indicated that corn's ability to sense urea was at least 7 orders of magnitude worse than that of the other three plants.Corn and pepper were cultivated under the condition that urea was the only exogenous nitrogen nutrient.The experimental results showed that at a concentration lower than the corn's ability to sense urea(1×10-10mol·L^-1),the corn died within 10-15 days and peppers could grow normally.It was proved that the linkage allosteric constant Ka obtained in this study reflected the true sensing ability of plant root-tip meristems to ureaand that corn's sensing of urea has some defects.