Project Number: 5428-21000-012-04
Project Type: Specific Cooperative Agreement

Start Date: Sep 01, 2005
End Date: Aug 31, 2010

Objective:
The objectives of this cooperative research project are to: 1) construct expressed sequence tag (EST) libraries from four normalized tissues of an interspecific hybrid of Pseudoroegneria, specifically Leymus cinereus X L. triticoides, of Pseudoroegnaria spicata, or bluebunch wheatgrass, Dactylis glomerata or orchardgrass, and Elymus, 2) array selected clones in microplates and sequence 10,000 clones from each of the two species, and 3) trim, filter, and annotate the sequences into a user-friendly bioinformatics pipeline, and release the sequencing results to the public via the in-house and NCBI websites.

Approach:
Four tissues will be collected for each of the four species. These tissues will be comprised of: rhizome and tiller buds, drought and salt stressed leaves, drought and salt stressed roots, and etiolated seedlings. Tissue will be harvested and immediately stored at -80 degrees C at Logan, UT. RNA will be extracted from the tissue using Trizol reagent, and purified through Qiagen RNeasy columns. Total RNA will be sent on dry ice to UIBC. Two normalized plasmid cDNA libraries will be prepared with 4 tissue specific 3' tags in each library. UIBC will pick and prep to sequence the 5' end of 9,984 clones per each of the two libraries. They will also direct bioinformatics on the sequence data including quality trimming, vector trimming, data storage, GenBank deposit, BLAST to comparative organisms, advanced analysis of ESTs including: clustering, assembly, annotation, and gene ontology associations.