柠檬醛致黄曲霉孢子丧失萌发力的机制

The Mechanism of Loss of Germination Ability of A.flavus Spore with Citral

通过由倒置显微镜、衍射光栅和线阵光电偶合器件CCD(chargecoupleddevice)等构成的显微多道分光光度系统及由计算机DEPHI编程工具编制的单细胞凝胶电泳SCGE(single cellgelelectro phoresis)图像分析系统 ,摄取荧光显微镜所呈图像 ,再由图像捕捉卡将CCD产生的图像信号送入计算机 ,将柠檬醛对黄曲霉质膜和核DNA损伤的图像进行显示存储和分析处理 ,测定彗星长度、荧光强度、矩类及头尾DNA含量比等彗星参数指标 .结果发现Olive尾矩、尾长、尾分布矩等彗星尾参数指标与柠檬醛致黄曲霉损伤浓度呈正相关性 ,当致损浓度达到 1 5mg L以上时 ,DNA损伤为致死性损伤 ,不能被细胞内修复系统所修复 .揭示柠檬醛通过损伤质膜而进入细胞 ,对DNA产生不可逆损伤 ,使孢子失去萌发力的机制 .实现将DNA损伤的生化定性检测推进到数值化研究范围 ,为柠檬醛的开发应用提供了重要理论依据 .与国内外同类技术相比 ,本检测观察系统还具有高灵敏度、快速、无扰、多光谱显微测定之特点 .

A fast multi-channel micro-spectrophotometer and a single-cell gel-electrophoresis imaging device were used for studying the mechanism of loss of germination ability of A.flavus spore with citral.The former device was composed of a inverted microscope,a differential grating and a linear array charge coupled device (CCD),while the latter was operated by a computer with a DEPHI program.A photo was taken under the fluorescence microscope via CCD and the image signal was transmitted to a computer via a frame grabber.The images of A.flavus plasma membrane and the DNA damage were displayed,stored,handled and analyzed.Four parameters,i.e.comet length,fluorescence intensity,moment and the ratio of head DNA% to tail DNA% were determined.It was found that there existed positive correlation between citral concentration and the above parameters.When the citral concentration reached above 1.5*!mg/L,DNA damage became lethal and not repairable in cell.It was also found that after citral entered into cell and damaged the plasma membrane,damaged DNA became irreversible with the loss of germination ability of the spore of A.flavus.Compared with other methods the method is rapid,sensitive,informative,quantitative and free of disturbance.