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Title: Construction and characterization of a half million clones BAC library of durum wheat (Triticum Turdigum SSP. durum).

Author
item CENCI, A - UC DAVIS, CA
item CHANTRET, N - UC DAVIS, CA
item KONG, X - USDA, ALBANY, CA
item Gu, Yong
item Anderson, Olin
item FAHIMA, T - UC DAVIS, CA
item DISTELFELD, A - UC DAVIS, CA
item DUBCOVSKY, JORGE - UC DAVIS, CA

Submitted to: Theoretical and Applied Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/14/2003
Publication Date: 6/27/2003
Citation: Cenci, A., Chantret, N., Kong, X., Gu, Y.Q., Anderson, O.D., Fahima, T., Distelfeld, A., Dubcovsky, J. 2003. Construction and Characterization of a half-million clones BAC library of durum wheat (Triticum Turdigum SSP. Durum). Theoretical and Applied Genetics. 107:931-939.

Interpretive Summary: Modern cultivated wheats come in two varieties, the durum wheat used for pasta-making and the bread wheats used for leavened and unleavened products such as raised and flat breads, cakes, cookies, etc. To continue to improve both types of wheats, the application of genomics technologies offers the greatest long-term opportunities. To fully apply genomics to wheat research and improvement, specific laboratory resources are needed, one of the most important of which is the construction of large cloned fragments of the wheat genomes. This report describes the generation of a library large collection of such clones for durum wheat. The library consists of 516,096 individual clones, each growing in an individual bacterial cell culture. The total of 1344 384-well plates represents enough of the total genome to expect to find any gene of interest with 94.4% probability. This library will serve as a resource for the entire wheat research community.

Technical Abstract: Durum wheat (Triticum turgidum ssp. durum, 2n=4x=28, genomes AB) is an economically important cereal used as the raw material for pasta and semolina making. In this paper we present the construction and characterization of a BAC library of tetraploid durum wheat cultivar 'Langdon'. This variety was selected because of the availability of substitution lines that facilitate the assignment of BACs to the A and B genome. The selected Langdon line has a 30 cM segment of chromosome 6BS from T. turgidum ssp. dicoccoides including a gene for high grain protein content that is the target of a positional cloning effort in our laboratory. A total of 516.096 clones were organized in 1344 384-well plates and blotted into 28 high-density filters. Ninety-eight percent of these clones have wheat inserts (0.3% chloroplast DNA, 1.4% empty clones and 0.3% empty wells). The average insert size of 500 randomly selected BAC clones was 131-kb, resulting in a coverage of 5.1 X genome equivalents for each of the two genomes, and a 99.4% probability of recovering any gene from each of the two genomes of durum wheat. Six known copy number probes were used to validate this theoretical coverage and gave an estimated coverage of 5.8 X genome equivalents. Screening of the library with eleven probes related to grain storage protein and starch biosynthesis showed that the library contains several clones for each of these genes, confirming the value of the library to characterize the organization of these important gene families. In addition, characterization of fingerprintings from colinear BACs from the A and B genomes showed a large differentiation between the A and B genomes. This library will be a useful tool for evolutionary studies in one of the best-characterized polyploid systems and a source of valuable genes for wheat. Clones and high-density filters can be requested at http://agronomy.ucdavis.edu/Dubcovsky/BAC-library/BAC_Langdon.htm.