Abstract The high-molecular-weight (HMW) subunits of wheat glutenin are the major determinants of the glu- ten visco-elasticity that allows wheat doughs to be used to make bread, pasta and other food products. In order to increase the proportions of the HMW subunits, and hence improve breadmaking performance, particle bom- bardment was used to transform tritordeum, a fertile amphiploid between wild barley and pasta wheat, with genes encoding two HMW glutenin subunits (1Ax1 and 1Dx5). Of the 13 independent transgenic lines recovered (a transformation frequency of 1.4%) six express the novel HMW subunits at levels similar to, or higher than, those of the endogenous subunits encoded on chromo- some 1B. Small-scale mixograph analysis of T 2 seeds from a line expressing the transgene for 1Dx5 indicated that the addition of novel HMW subunits can result in significant improvements in dough strength and stability, thus demonstrating that transformation can be used to modify the functional properties of tritordeum for im- proved breadmaking. Key words Tritordeum · Transformation · HMW glutenin subunits · Seed protein engineering · Dough functionality Introduction Tetraploid pasta wheat (Triticum turgidum conv. durum Dest. em.M.K., genome constitution AABB) is well adapted to the hot dry Mediterranean climate of southern Europe and North Africa, where it is traditionally used to make pasta, flat breads, bulgar and couscous (Matsuo 1996). It is not well suited for making leavened bread, lacking the D genome associated with high-gluten visco- elasticity present in hexaploid bread wheat (Triticum aestivum L., AABBDD), although significant amounts are used for this purpose in southern Italy and Greece (Matsuo 1996). Tritordeum (x Tritordeum Ascherson et Graebner) is an amphiploid derived from crossing the South American wild barley Hordeum chilense Roem et Schulz (H ch H ch ) with either pasta wheat or bread wheat, resulting in hexaploid and octaploid forms, respectively (Martin and Chapman 1977; Martin and Sanchez-Monge Laguna 1982; Martin et al. 1996). Tritordeums have been shown to have good agronom- ic performances and similar yields and protein contents to the parental wheat varieties (Ballesteros 1993; Alva- rez et al. 1992). However, the grain quality is generally inferior to those of the wheat parents (Alvarez et al. 1994) and none of the hexaploid tritordeums so far pro- duced are suitable for commercial bread production. Genetic transformation of tritordeum has recently been achieved, by particle bombardment of immature in- florescences (Barcelo et al. 1994; Barcelo and Lazzeri 1995). This provides an opportunity to improve the breadmaking performance of tritordeum by the introduc- tion of gluten protein genes that are known to be associ- ated with breadmaking quality. One group of gluten pro- teins is of particular interest in this respect as they ap- pear to be major determinants of gluten visco-elasticity and hence breadmaking quality. These are the high-mo- lecular-weight (HMW) subunits of wheat glutenin. Two HMW subunit genes are present on each group-1 chro- mosome, encoding a high M r x-type and a low M r y-type subunit. However, variation in the expression of these genes results in differences in the number of proteins Communicated by J.W. Snape L. Rooke · S. Steele · P.A. Lazzeri · P. Barcelo ( ✉ ) IACR-Rothamsted, Harpenden, Herts, AL5 2JQ, UK e-mail: pilar.barcelo@gbr.dupont.com Fax: +44 1582 768791 F. Barro · A. Martin Instituto de Agricultura, Apdo 4084, Cordoba 14080, Spain A.S. Tatham, R. Fido, P.R. Shewry IACR-Long Ashton, Department of Agricultural Sciences, University of Bristol, Long Ashton, Bristol, BS18 9AF, UK F. Békés · P. Gras Plant Science CRC, CSIRO, North Ryde, NSW 2113, Australia Theor Appl Genet (1999) 99:851–858 © Springer-Verlag 1999 ORIGINAL PAPER L. Rooke · F. Barro · A.S. Tatham · R. Fido · S. Steele F. Békés · P. Gras · A. Martin · P.A. Lazzeri P.R. Shewry · P. Barcelo Altered functional properties of tritordeum by transformation with HMW glutenin subunit genes Received: 15 January 1999 / Accepted: 5 February 1999