Brachypodium distachyon has recently been established as a model for cereals and temperate grasses because of its sequenced small genome (~270M), a small stature, short life cycle, self-fertility, an efficient transformation system and particularly its diverse ecotypes. Determination of the function for each gene in the Brachypodium genome is urgently needed for utilization of this species. In this study we (1) analyze the expression of each gene involved in vernalization pathway in Brachypodium and the correlation of vernalization requirement with freezing tolerance, (2) develop a novel high-throughput RNAi method, Phi29-Amplified RNAi Construct (PARC), to construct a cold-specific RNA interference library, for detecting the mechanism of cold acclimation and how plants develop freezing tolerance in Brachypodium, (3) knockdown brassinosteroid receptor gene BRI1 by PARC in Brachypodium to characterize the loss-of-function BdBRI1-RNAi mutants. These studies will shed light on the pathways involved in vernalization, cold acclimation, freezing tolerance, brassinosteroids signaling, and drought tolerance. PARC can either be use to generate a high-throughput long hairpin (lhRNA) expression library or a lhRNA construct of a single gene.