January 15-19, 2011
Town & Country Convention Center
San Diego, CA
Marília Pappas1 , Alessandra Reis2 , Giancarlo Pasquali3 , Georgios Pappas Jr.1,2 , Dario Grattapaglia1,2
Micro RNAs are a class of small non-coding RNAs that are key regulators of gene expression. Micro RNAs are involved in a wide variety of physiological processes in plants such as development, flowering and response to stress. As part of the ongoing Eucalyptus grandis genome sequencing consortium (Eucagen), we set to provide experimental support for the characterization of the small RNA fraction of this genus and to help the annotation process. We also wanted to probe for interspecific differences that could account for the marked difference in wood quality between E. grandis and E. globulus. Total RNA was extracted from xylem of two genotypes of E. globulus, and xylem and leaves of one genotype of E. grandis - BRASUZ, the genotype being sequenced by JGI. The samples were subjected to deep sequencing using one lane of a Illumina GAII. Bioinformatics processing revealed that all samples had the expected bimodal distribution peaking at 21 and 24 bp. The 24 bp fraction was consistently more abundant than the 21 bp fraction, and the normalized counts ratio between these two showed significant differences between tissues and species. Following miRNA annotation standards, the 21 bp sequences were mapped against E. grandis 8X draft sequence and checked for compatible secondary structure of the putative precursors. The filtered set of miRNA candidates revealed several highly conserved families, present in miRBase, but also a significant amount of Eucalyptus specific miRNA families. Validation experiments via RT-PCR are in progress for a set of both categories.