Small RNAs ranging in size between 20 and 32 nt regulate gene expression through chromatin modification, mRNA degradation, and translational repression. Three major classes of small RNAs have been characterized: microRNAs (miRNAs), short interfering RNAs (siRNAs), and Piwi-interacting RNAs (piRNAs). miRNAs are expressed in a developmentally regulated and tissue-specific manner and are involved in development and cell differentiation. siRNAs are mainly involved in defense against transposons and viruses. piRNAs are expressed in germ cells and stem cells and are thought to repress transposition of retrotransposons. In this chapter, we describe the methods of small RNA cloning, annotation and classification, and their expression analysis during development.