洋葱表皮和小麦幼苗自发荧光的研究

Study on Autofluorescence of Onion Epidermis (Allium cepa) and Wheat Seedlings (Triticum aestivum)

摘要: 自发荧光是多数植物组织具有的特征，这对于通过荧光标记手段研究基因表达非常不利，对自发荧光的认识有助于对其进行科学地应用。本研究选择常用的洋葱表皮细胞做参照，对比研究小麦的自发荧光现象，将小麦在有光照、无光照下幼苗不同发育天数的根、叶鞘、叶片制成装片，在激发光光源的作用下，观察小麦不同时期根、叶鞘、叶片的蓝色、绿色、红色自发荧光的分布情况；最后通过50 μg∙L⁻¹的卡那霉素处理使小麦幼苗白化，制备叶片装片观察三种颜色的荧光。实验结果表明黄皮洋葱的内、外表皮细胞均检测不到红、绿、蓝色荧光；紫皮洋葱的表皮细胞色素会影响红色荧光的观察，也不具有荧光。在500 nm波长的激发光照射下，见光小麦的根在第1 d以及6~10 d具有红色荧光；叶片及叶鞘从第3 d起一直有强烈的红色荧光；黑暗培养的小麦根在第16 d出现红色荧光；叶鞘始终无荧光产生，叶片有较弱的红色荧光。450 nm激发光照射下，见光小麦，根在第1 d以及6~10 d有明显的绿色荧光；黑暗培养的小麦根从第16 d起出现绿色荧光。光照、黑暗下培养的小麦叶鞘都从第22 d起有明显的绿色荧光，两种条件下培养的小麦叶片始终检测不到绿色荧光；400 nm激发光照射下，小麦的根、叶鞘、叶片中蓝色荧光始终存在，且叶鞘中蓝色荧光随着发育天数增加逐渐强烈；处理后白化的叶片无法观察到红色和绿色荧光，蓝色荧光依然存在。白化处理后有利于通过红色荧光蛋白标记目的基因，为小麦的组织、细胞定位研究提供便利。

Abstract: Autofluorescence is a characteristic of most plant tissues, which is very unfavorable for studying gene expression through fluorescent labeling methods. The understanding of plant autofluorescence can benefit for proper application. In this study, the commonly used onion epidermal cells were selected as a reference to compare and study the autofluorescence phenomenon of wheat seedling. The roots, sheaths, and leaves of wheat seedlings in different days of development under light and dark were used to make slices to stimulate the light source. Next, we observed the distribution of blue, green, and red autofluorescence in the roots, leaf sheaths, and leaves of wheat in different periods. Finally, the wheat seedlings were albino treated with 50 μg∙L⁻¹ kanamycin, and the leaves were mounted to observe the fluorescence of three colors. The results of the experiment showed that the red, green, and blue fluorescence could not be detected in the epidermal cells of the yellow onion; the epidermal cytochrome of the purple onion would affect the observation of red fluorescence, and it did not have fluorescence. Under the excitation light of 500 nm wavelength, the roots of visible wheat have red fluorescence on the 1st and 6~10 d; the leaves and leaf sheaths have always had strong red fluorescence from the 3rd day onwards; the wheat roots in the dark have red fluorescence on the 16th day; The leaf sheath always has no fluorescence, and the leaf has weak red fluorescence. Under 450 nm excitation light, the roots showed obvious green fluorescence on the 1st day and 6~10 d when the wheat was exposed to light; in the dark, the roots showed green fluorescence on the 16th day. The leaf sheaths of wheat cultivated under light and dark showed obvious green fluorescence on the 22nd day, and the leaves of wheat cultivated under the two conditions could not always detect green fluorescence; under the irradiation of 400 nm excitation light, in the roots, sheaths, and leaves of wheat, the blue fluorescence always exists, and the blue fluorescence in the leaf sheath becomes stronger with the increase of the development days; the red and green fluorescence cannot be observed in the albino leaves after treatment, and the blue fluorescence still exists. After bleaching treatment, red fluorescent protein can be used to label the target gene, which provides convenience for the study of the tissue and cell location of wheat.