制片方法对枣花授粉荧光显微观察效果的影响

Slicing methods for fluorescence microscopic observation of style of Chinese jujube after pollination

【目的】探究适合观察枣花授粉荧光显微观察效果的最佳制片方法,为枣树授粉受精研究提供方法依据。【方法】以4年生、生长发育良好的‘中秋酥脆枣’枣花作为试验材料,经过固定、水洗等步骤,采用不同软化和染色方法对枣花进行处理,压片后在荧光显微镜下拍照,比较不同处理下枣花授粉的荧光显微观察效果。采用NaOH溶液对枣花柱进行软化,设置1、2、3、4和8 mol/L 5个浓度梯度,分别软化0.5 h和1 h,软化后用水溶性苯胺蓝溶液对枣花进行染色,设置0.01%、0.05%和0.10%3个浓度梯度,分别染色0.5、1和4 h。【结果】相同处理时间,NaOH溶液浓度越高,枣花花柱软化程度也越高;而NaOH溶液浓度越高,所需要的软化处理时间也越短。选择1、2和3 mol/L NaOH溶液对枣花进行软化,都能取得良好的压片效果,但当NaOH溶液浓度超过4 mol/L后,枣花花柱组织明显变软,压片时组织也容易破碎。在相同软化方法处理的条件下,同一浓度水溶性苯胺蓝溶液的染色时间增加,对花粉管的染色效果并没有产生明显的增强作用;而在相同染色时间内,用高浓度的水溶性苯胺蓝溶液对枣花进行染色,染色效果较低浓度的要好。【结论】1)选择1 mol/L或2 mol/L NaOH溶液来处理枣花花柱,都能取得较好的制片效果。2)在软化时间的选择上,软化0.5 h就能取得较好的压片效果,这能缩短试验需要的时间;在相同染色时间内,用高浓度的水溶性苯胺蓝溶液对枣花进行染色,染色效果较低浓度的要好。

【Objective】In order to explore the best slice production of microscopic observation by fluorescent method for the Stigma and Style,and provide some study basis for the pollination and fertilization of Chinese jujube,the flowers for‘Zhongqiusucui’,which grew four years and was well-developed,were selected as the experimental material.【Method】After fixation and rinsing in distilled water,these flowers by processing with different softening and dyeing methods,were squashed and observed under OLYMPUS BX51 fluorescent microscope.Then pictures were taken,which were used to compare different effects of fluorescence observation processed with different methods.The jujube flowers were treated in 1 mol/L,2 mol/L,3 mol/L,4 mol/L and 8 mol/L NaOH for 0.5 h and 1 h respectively to soften the tissue.Staining was accomplished next in 0.01%,0.05% and 0.1% solution of water-soluble aniline blue for 0.5 h,1 h and 4 h respectively.【Results】At the same treatment time,the higher the concentration of NaOH,the higher the softening degree of jujube flowers is.The higher the concentration of NaOH solution,the shorter the softening time.After more than 4 mol/L NaOH,the tissue of jujube flower becomes soft obviously and easy to be squashed.Choosing 1 mol/L,2 mol/L or 3 mol/L NaOH,better effect of softening can be achieved.For the same softening method,when the dyeing time of the water-soluble aniline blue solution with the same concentration increases,the dyeing effect on the pollen tube is not significantly enhanced.For the same dyeing time,with high concentration of water-soluble aniline blue solution dyeing jujube flowers,dyeing effect is better than low concentration.【Conclusion】1)Choosing 1 mol/L or 2 mol/L NaOH,better effect of softening can be achieved.2)For time,softening 0.5 h can result in a better effect;for the same dyeing time,with high concentration of water-soluble aniline blue solution dyeing jujube flowers,dyeing effect is better than low concentration.