Figure 1.

Outline of the experimental protocol used for preparation of small RNA libraries. RNA was isolated from ZF developmental samples and from adult tissues using TRIzol^® and fractionated on 12.5% denaturing PAGE. Small RNAs were purified from those gels and then ligated to a 3' adapter (AMP-5'p-5'p/CTGTAGGCACCATCAATdi-deoxyC- 3') and to a 5' linker (see Methods). cDNA was prepared and amplified using 20 PCR cycles. PCR products were subjected to clonal amplification by emulsion PCR and then pyrosequenced using a 454 genome sequencer.