Bio-Cell Gene Expression Analyzer

术语“生物细胞”是指在整个应力施加实验中保持其存活状态的细胞。进行这项研究是为了证明生物细胞对各种应力信号的响应的真实的时间分析。将电应力信号施加到烟草或水稻的培养细胞。作为细胞应答，用显微荧光图像分析系统测量基因表达。本文以烟草BY-2细胞为实验材料，在不同强度的脉冲电应力信号作用下，测定了细胞膜的电阻抗。结果表明，当跨膜电位（V_）超过200 mV时，细胞膜发生了明显的透化作用<CMP>。同时，利用水稻原生质体进行的两步实验证实了几丁质酶基因的表达依赖于Ca^&lt;2+&gt;。在第一步中，Ca^2+的引入由脉冲电应力信号控制。第二步，我们通过直接向原生质体中注入Ca^&lt;2 +&gt;来证明几丁质酶基因的表达。除了电应力之外，我们还打算在生物细胞内部施加电化学应力。在此基础上，我们研制了直径为1-2 μm的毛细管碳微电极，并通过对多巴胺的循环伏安曲线的分析来考察其性能。综上所述，本研究成功开发了生物细胞基因表达分析系统，并展示了其良好的性能。

The term "bio-cell" means a cell that maintains its viable state throughout the experiments of stress application. This study was done in order to demonstrate the real time analysis of the bio-cell responses to various stress signals. An electric stress signal was applied to cultured cells of tobacco or rice. As the cell response, the gene expression was measured with a microscopic fluorescent image analyzing system. By the experiment using tobacco cells, line BY-2, the electric impedance of the cell membrane was measured under various intensities of pulsing electric stress signals. As the result, appreciable membrane permeabilization was observed when the cross membrane potential (V_<CMP>) exceeded 200mV.On the other hand, the Ca^<2+>-dependent expression of chitinase gene was demonstrated by 2-step experiments using rice protoplasts. At the first step, Ca^<2+> introduction was controlled by the pulsing electric stress signal. As the result, Ca^<2+> entered the cells around 1-5 mV of VCMP.As the 2nd step, we demonstrated the chitinase gene expression by the direct injection of Ca^<2+> into the protoplasts. Besides the electric stress, we intended to apply an electrochemical stress to a bio-cell at its inside. Then we developed a capillary carbon microelectrode with a 1-2 μm diameter and checked its performance by analyzing cyclicvoltammograms of dopamine. In conclusion, the analyzing system of the gene expression of a bio-cell was developed and its promising performance was demonstrated.