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胡萝卜包埋制作拟南芥徒手切片

庄严 |  2020-12-29  | DOI: 10.21769/BioProtoc.v621
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视频简介 本文描述了一种简便的胡萝卜包埋方法,无需任何仪器辅助,极大地降低拟南芥花茎徒手切片的操作难度,可以对各种突变体快速、大量地获得拟南芥组织切片,观察木质素的沉积状态。
  • 视频介绍

实验要点:

我亦无他,唯手熟尔!Practice makes perfect!

 

实验材料:

拟南芥花茎、胡萝卜、1号毛刷、吉列刀片、尖头镊子、美工刀、盖玻片、载玻片、200微升移液器、擦手纸、自来水

 

实验步骤:

  1. 用美工刀将胡萝卜切成柱形,保持上表面水平。Cut the carrot into a column or prism with a knife. Keep the upper surface flat and horizontal. 
  2. 用尖镊子在柱上戳一个小孔,小孔的直径近似于拟南芥花茎。Punch a hole on the prism with a sharp forcep. The diameter of the hole is similar to that of Arabidopsis stems.
  3. 将花茎插入小孔。Insert one stem into the hole slightly with a hand or forcep.

  4. 切除伸出在外面的多余花茎,确保胡萝卜小孔内有约0.5cm长的花茎段。Cut the stem that stretches out of the hole. Make sure the stem inside the hole about 0.5cm long.
  5. 一只手握住胡萝卜,保持上表面水平,用吉列刀片水平切出一张薄片,切片尽量薄而不破。Hold the carrot with a hand and keep the upper surface horizontal. Cut a thin section with a razorblade (Gillette). Make the section as thin as possible but intact.

  6. 切片会停留在刀片上,用半湿的1号毛刷轻轻将花茎切片挑出,置于含有200uL水的载玻片上。The section will be on the razorblade after cutting. Move the section with a half-wet painting brush slightly into 200uL water on the slide. 

  7. 盖好盖玻片,在显微镜下观察切片。可以预先用间苯三酚对木质素染色(下图左),或直接在紫外光下观察木质素自发荧光(下图右)。Cover the cover glass. Examine the section under the microscope. Lignin is stained by phloroglucinol (left) or observed by self-fluorescence under UV light (right).

 

[关键词] 拟南芥、花茎、徒手切片、木质素
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