【作者】 贾莉君；

【导师】 沈其荣；

【作者基本信息】 南京农业大学 ， 植物营养学， 2006， 硕士

【摘要】 离子选择性微电极是一种在活体条件下，测定某一单个细胞内离子的活度和细胞内离子活度的动态变化，与其它测定方法相比，具有连续测定、记录瞬时变化和真实可靠的特点。不仅如此，利用微电极技术还可以测到一个细胞内不同细胞器内离子的活度。迄今为止，国外研究主要集中在利用微电极技术对大麦、玉米、大豆等作物根系养分离子的研究。由于水稻根系容易木质化，根系坚硬测定过程中易造成对微电极的严重破坏，加大了研究的难度，所以关于微电极技术在水稻这一中国主要的粮食作物根系养分离子研究中的甚少。植物叶片较薄，测定时容易发生震动使得微电极易离开细胞，很难得到稳定的测定数据，国内外相关的研究尚处于空白。因此，本文着力于开发微电极技术测定植物叶片的方法以及其在水稻根系硝态氮的吸收和利用方面的研究。 首先，通过多次重复的实验和摸索，在国内首次详细介绍了用微电极技术研究植物细胞电生理性质的方法，包括电生理实验系统的组建，单孔及双阻离子选择性微电极的制作方法、工作原理、根系和叶片细胞中离子活度测定方法及注意事项等。 实验证明双阻硝酸根离子选择微电极对细胞中硝酸根离子有较低的检出限，是一种选择性高、灵敏、经济的活体测定植物细胞中离子活度的方法。用微电极测定发现用含有5mmol·L^-1 NO₃^-的营养液诱导48h后的六叶期小白菜，叶片细胞中硝酸根离子活度分布在活度高低明显不同的两个区间内：细胞质中是0.24～10mmol·L^-1，液泡中20～110mmol·L^-1，且两个区间在细胞跨膜电位上也有差异。液泡占整个细胞体积的90％，所以，植物所吸收的硝酸根离子都集中在液泡中。 其次，运用单孔微电极插入法研究了在不同细胞内外pH值的条件下，水稻——常规籼稻（杨稻6号）吸收NO₃^-时细胞膜电位的响应特征。结果表明：（1） 大多数情况下，未经NO₃^-培养的水稻根表皮细胞吸收NO₃^-会引起细胞膜电位瞬时的去极化，偶尔会产生膜电位的超级化。当水稻根系吸收NO₃^-时，表皮细胞膜电位改变的程度随外界NO₃^-浓度的增大而增大，在浓度为0.5mmol·L^-1时达到最大；（2） 在不同外界pH更多还原

【Abstract】 The ion selective microelectrode is measuring ion activity in a single cell in vivo and its continuous change during external treatment. It is characteristic of continuous measuring, reliable and without cell fraction. Ion activity in different compartmentations of cell can also be measured using ion selective microelectrode. Many reports have been found on the root ion uptake of maize, barley, and soybean, etc. Unfortunately, rice roots can be easily ligilified, thus, there were only a few reports concerning root ion uptake of rice which servers the main food in China. Plant leaf was so thin and easily vabntated that it was hard to matain a steady recording, the using of microelectrode studying ion uptake on plant leaf cell is left as a blank. Hence, the thesis was set out to studying the using of ion selective microelectrode in rice leaf and extends its application in studying nitrate uptake and utilization in rice root.Firstly, ion selective microelectrode and its application in nitrate uptake were firstly introduced in China. The protocol of manipulation was described in detail after practice of several times, including the construction of electrophysiology lab, making of single and double barreled selective microelectrode, working theory and notice.The microelectrode was turn out to be characterized by a high selectivity and sensitivity of ion and a good economy compared with other intracellular measurement. In addition, the in vivo method is better than the chemical analysis of the samples. The detected results showed that nitrate activity in leaf cell of Chinese cabbage induced with 5 mmol·L^-1 nitrate for 48h fell into two different populations, i.e.0.24 mmol·L^-1-10 mmol·L^-1 in cytosol and 20 mmol·L^-1-110 mmol·L^-1 in vacuole. Since vacuolar volume accounts for 90% of a whole cell, most of the nitrate in plants is stored in vacuoles. And, thus it is of great importance to study the remobilization of the nitrate in vacuoles both in increasing nitrogen use efficiency and in decreasing the nitrate content the vegetable. Then, the uptake of nitrate (NO₃^- by roots of rice seedlings （yangdao 6） at different pHs更多还原