A new method for rapid visualization of defects in leaf cuticle reveals five intrinsic patterns of surface defects in Arabidopsis
A new method for rapid visualization of defects in leaf cuticle reveals five intrinsic patterns of surface defects in Arabidopsis
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DOI：
10.1046/j.1365-313x.2003.01946.x
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发表时间：
2004-01-01
期刊：
影响因子：
7.2
通讯作者：
Machida, Y
中科院分区：
文献类型：
--
作者：
Tanaka, T;Tanaka, H;Machida, Y
高等植物的表皮产生角质层，覆盖每株植物的外表面。角质层在植物发育过程中起着重要作用，一些角质层缺陷的突变体表现出形态异常，如器官融合。角质层形成的方式及其对形态发生的贡献知之甚少。传统的检测角质层的透射电子显微镜（TEM）需要费力的程序，其中包括固定，染色与锇，超薄切片的准备。由于可以检查的标本的大小有限，也很难调查展开的叶子的整个表面。因此，TEM不适合于大规模筛选角质层缺陷突变体。我们在这里描述了一种快速，廉价的方法，指定的甲苯胺蓝（TB）测试，检测角质层缺陷在整个叶片。我们证明了TB测试的有效性，使用突变体的拟南芥，包括异常叶形状1（ale 1），fiddlehead（fdh），和5 eceriferum（cer）突变体，其中的结构和/或功能的角质层是异常的。使用TB测试对突变体进行遗传筛选使我们能够识别7个位点。突变体叶片的角质层缺陷区域揭示了五种内在的表面缺陷模式（I类至V类），表明叶片上功能性角质层的形成涉及各种空间调控因素。
The epidermis of higher plants generates the cuticle layer that covers the outer surface of each plant. The cuticle plays a crucial role in plant development, and some mutants with defective cuticle exhibit morphological abnormalities, such as the fusion of organs. The way in which the cuticle forms and its contribution to morphogenesis are poorly understood. Conventional detection of the cuticle by transmission electron microscopy (TEM) requires laborious procedures, which include fixation, staining with osmium, and preparation of ultra-thin sections. It is also difficult to survey entire surfaces of expanded leaves because of the limited size of specimens that can be examined. Thus, TEM is unsuitable for large-scale screening for mutants with defective cuticle. We describe here a rapid and inexpensive method, designated the toluidine-blue (TB) test, for detection of cuticular defects in whole leaves. We demonstrated the validity of the TB test using mutants of Arabidopsis thaliana, including abnormal leaf shape1 (ale1), fiddlehead (fdh), and five eceriferum (cer) mutants, in which the structure and/or function of the cuticle is abnormal. Genetic screening for mutants using the TB test allowed us to identify seven loci. The cuticle-defective regions of leaves of the mutants revealed five intrinsic patterns of surface defects (classes I through V), suggesting that formation of functional cuticle on leaves involves various spatially regulated factors.