Funct Integr Genomics (2003) 3:153–159 DOI 10.1007/s10142-003-0092-8 ORIGINAL PAPER Dipak K. Santra · Devinder Sandhu · Thomas Tai · Madan K. Bhattacharyya Construction and characterization of a soybean yeast artificial chromosome library and identification of clones for the Rps6 region Received: 9 October 2002 / Revised: 21 June 2003 / Accepted: 6 August 2003 / Published online: 1 October 2003  Springer-Verlag 2003 Abstract We report the construction and characterization of the first soybean yeast artificial chromosome (YAC) library using high-molecular weight DNA isolated from leaf nuclei of the cultivar Conrad 94 that carries Phytophthora resistance genes Rps1-k and Rps6. The quality of this library has been evaluated through analysis of 393 randomly selected YAC clones. The library consists of 36,864 clones, of which 19,956 carry single soybean YACs with an average size of about 285 kb. The library represents approximately five soybean genome equivalents. The probability of finding any soybean sequences from this library is about 0.99. The library was screened for 43 SSR markers representing the whole soybean genome. We were able to identify positive YAC pools for 95% of the SSR markers. Two YAC clones carrying molecular markers linked to the Rps6 gene were identified. The YAC library reported here would be a useful resource for map-based cloning of agronomically important soybean genes and also to complement the effort towards construction of the physical map for the soybean genome. Keywords Soybean · YAC library · Positional cloning · Resistance genes Introduction Large insert genomic libraries such as yeast artificial chromosome (YAC) libraries have been constructed for many important plant species like Arabidopsis (Grill and Somerville 1991), rice (Umehara et al. 1995), corn (Edwards et al. 1992), barley (Kleine et al.1993), tomato (Martin et al. 1992), potato (Leister et al. 1997) and pepper (Tai and Staskawicz 2000). Such libraries have facilitated positional cloning of genes and genomic regions in many of these species (Martin et al. 1993; Bent et al. 1994; Yoshimura et al. 1996; Brommon- schenkel and Tanksley 1997; Tai et al. 1999; Teraishi et al. 2001), physical genome mapping (Kurata et al. 1997; Sato et al. 1998; Saji et al. 2001), analysis of repetitive sequences (Dunford and Rogner 1991; Schmidt et al. 1994), and studying the relationship between genetic and physical distances (Civardi et al. 1994). The main advantages of YACs over prokaryotic-based cloning systems are its large insert size (100–1000 kb), and ability to maintain sequences that are unstable and not well represented in prokaryotic cloning system. Nearly all DNA sequences are clonable in YACs (Schlessinger et al. 1991), as yeast can tolerate several kinds of DNA structures that may not be properly propagated in Escherichia coli (Burke et al. 1987). The power and utility of YAC libraries is particularly evident from positional cloning of agronomically important genes with a reduced number of chromosomal walking steps from crop species that have large and complex genomes (e.g. Martin et al. 1993; Tai et al. 1999). Soybean, one of the most important crops in North America, is a diploidized tetraploid (Hadley and Hy- mowitz 1973; Shoemaker et al. 1996). Several soybean BAC libraries have been constructed (Marek and Shoe- maker 1997; Danesh et al. 1998; Salimath and Bhat- tacharyya 1999; Tomkins et al. 1999; Meksem et al. 2001) and are being used for physical mapping of the soybean genome (Marek et al. 2001; Meksem et al. 2001). The average insert sizes in these libraries are reported to be about 100–150 kb. To date no usable soybean YAC D. K. Santra · D. Sandhu · M. K. Bhattacharyya ( ) ) Department of Agronomy, Iowa State University, Ames, IA 50011, USA e-mail: mbhattac@iastate.edu T. Tai USDA-ARS, Agronomy and Range Science, University of California, One Shields Ave, Davis, CA 95616-8515, USA Present address: D. K. Santra, Department of Crop and Soil Sciences, Washington State University, Pullman, WA 99164-6420, USA