Stern David L, Sucena Elio
Cold Spring Harb Protoc. 2011 Nov 1;2011(11):1394-8. doi: 10.1101/pdb.prot066498.
The Drosophila cuticle carries a rich array of morphological details. Thus, cuticle examination has had a central role in the history of genetics. Studies of the Drosophila cuticle have focused mainly on first-instar larvae and adult cuticular morphology. Although the cuticles of second- and third-instar larvae are strikingly different from those of the first instar, these differences have been poorly studied. This protocol describes three methods for preparing cuticles from fed larvae. One commonly used procedure involves manually pricking the larvae. A simpler method for preparing larval cuticles is to burst the larvae once they have been mounted. This method is used for first- and second-instar larvae and does not require pricking; it removes the gut contents by "popping" the rear of the embryo using pressure from the coverslip. If just the right amount of medium is used, the coverslip will be pulled toward the slide, applying pressure on the samples. The larvae usually burst from their posterior ends. Also presented is an alternative procedure designed specifically for the use with third-instar larvae, although the "pricking" method can be used at this stage.
果蝇的表皮具有丰富多样的形态细节。因此，表皮检查在遗传学历史中一直占据核心地位。对果蝇表皮的研究主要集中在一龄幼虫和成虫的表皮形态上。尽管二龄和三龄幼虫的表皮与一龄幼虫的表皮有显著差异，但这些差异尚未得到充分研究。本方案描述了三种从取食后的幼虫制备表皮的方法。一种常用的方法是手动刺破幼虫。制备幼虫表皮的一种更简单的方法是在幼虫固定后将其弄破。这种方法适用于一龄和二龄幼虫，不需要刺破；通过用盖玻片施加压力“弹出”胚胎后端来去除肠道内容物。如果使用适量的培养基，盖玻片会被拉向载玻片，对样本施加压力。幼虫通常从后端破裂。还介绍了一种专门为三龄幼虫设计的替代方法，不过在这个阶段也可以使用“刺破”方法。
