Monday, November 5, 2007
81-2

Construction of a BAC Library of the Leymus cinereus X L. triticoides Hybrid.

Steve Larson1, Chantel Scheuring2, Bradley Bushman3, Ivan W. Mott1, Jianmin J. Dong2, Yang Zhang2, Xiaojun Zhang2, Hong-Bin Zhang2, and Richard Wang4. (1) USDA-ARS, USDA-ARS Forage & Range Research Laboratory, Utah State University, Logan, UT 84322-6300, (2) Texas A & M University, College Station, TX 77843-2474, (3) USDA-ARS-NPA-SPNRU, USDA-ARS Forage and Range Research Lab, 695 N 1100 E, Logan, UT 84322-6300, (4) 695 North 1100 East, USDA-ARS, USDA-ARS, Forage & Range Research Laboratory, Logan, UT 84322-6300

Leymus cinereus and L. triticoides are two wildrye grasses with many contrasting morphological and agronomic traits. The interspecific hybrid Leymus cinereus X L. triticoides and its progenies had been used to develop linkage maps of traits and molecular markers. To generate genomic DNA sequence information for these important wildrye species, we are developing a bacterial artificial chromosome (BAC) library with an average insert size of 130 kb or larger for this interspecific hybrid. The DNA was partially digested with two restriction enzymes, BamHI and EcoRI, and then ligated into two vectors of different systems -- F plasmid-based BAC vector (pECBAC1) and P1 plasmid-based plant-transformation-competent binary BIBAC or TAC vector (pCLD04541). The combined BAC and BIBAC ligations yielded approximately 406,000 clones, which were preserved in 1057 384-well micro-plates. It was estimated that the BIBAC clones had an average insert size of 135 kb and the BAC clones had an average insert size of 155 kb. The cloned DNAs arrayed on 22 filters (22 x 22 cm each) represent 3x coverage of the Leymus genome, and will be screened with several sets of overgo primers to identify BAC clones containing candidate genes for important agronomic traits.