Ruby Orange Developing Flower cDNA Library

Vector: Lambda Uni-ZAP XR, excised phagemid; Site_1: EcoRI; Site_2: XhoI; Two trees with open-pollinated flowers in a mixed planting at UC Riverside Agricultural Operations were the source of tissue. Flower buds were collected by Federici (Roose lab) from January March 2003 in four groups: 1) no white petals, 2) white petals visible, bud less than 0.5 cm, 3) bud more than 0.5 cm but not open, 4) newly opened flowers. Tissues were snap frozen and then stored at -80C until further processing. Fenton (Close lab) purified RNA by the phenol method described in J. Japanese Soc. Hort. Sci. 1996. 64 (4): 809-814, purified poly(A) mRNA using a PolyATtrack mRNA Isolation System IV (Promega), produced a primary cDNA library using a lambda ZAP XR cDNA Synthesis Kit (Stratagene), then mass-excised one million pfu from the primary library to produce a phagemid population. Phagemids were plated, plasmid DNA purified, cDNA clones archived, and DNA sequences determined bi-directionally using an ABI3730 at the Arizona Genomics Institute, University of Arizona (Kim, Kudrna, Stum, Yost, Wing). Chromatogram files were downloaded by FTP to UC Riverside (by Close), then processed at UC Riverside (by Wanamaker, Close lab) using the HarvEST pipeline (http://harvest.ucr.edu) to remove vector and cloning oligo sequences and various contaminants, and to trim to a high quality region. Sequences that retained a phred 17 region of at least 100 bases were deposited to GenBank.