454 sequencing of Eucalyptus grandis transcript fragment library
Three greenhouse-grown Eucalyptus grandis seedlings from each of seven open-pollinated families (21 genotypes in total) were dissected into xylem (Xy), phloem (Ph), roots (R), young leaves(YL), mature leaves (ML) and apical/lateral meristems (M). Tissues were immediately frozen in liquid nitrogen and stored. For each tissue, RNA was extracted from a pool of equal proportion of plant material from all 21 genotypes. RNA isolated from each tissue pool was combined in varying proportions (10% YL, 10% ML, 15% R, 15% Ph, 20% M and 30% Xy) to a single pool. cDNA was synthesized with Clontech SMART cDNA Library Construction Kit (Qiagen USA, Valencia, CA) and normalized with Evrogen Trimmer-Direct Kit (Evrogen, Moscow).