**1. Introduction**

Narrow genetic diversity often limits the improvement of many traits in wheat. The introgression of genes from wild relatives to wheat has become a widely recognized genetic approach for increasing genetic diversity and, hence, the need to explore primary, secondary and tertiary gene pools of wheat has grown [1,2]. *Hordeum chilense* Roem. et Schultz. is a diploid wild barley that exhibits advantageous agronomic and quality characteristics [3–6]. Furthermore, its high crossability with other species of the tribe Triticeae, such as both durum and common wheat, [3–5] make it useful in cereal breeding.

Addition and substitution lines of alien chromosomes in a common wheat background, are useful for introgressing alien chromosomal segments carrying genes of agronomical interest into wheat. Chromosome addition lines of *H. chilense* in the *Triticum aestivum* L. cultivar "Chinese Spring" have been obtained including five for chromosomes 1Hch, 4Hch, 5Hch, 6Hc<sup>h</sup> and 7Hc<sup>h</sup> and the ditelosomic addition line for the short arm of chromosome 2Hc<sup>h</sup> [7]. Fertile wheat lines carrying deletions and translocations

involving chromosome 3Hc<sup>h</sup> from *H. chilense* have also been obtained [8]. However, no addition or substitution lines in a common wheat background have been developed for chromosome 2Hch.

The location of agronomic traits on specific *H. chilense* chromosomes have been carried using these available wheat—*H. chilense* addition and substitution lines, such as resistance to greenbug (*Schizaphis graminum* Rond.) [9] and endosperm prolamins located on chromosome 1Hc<sup>h</sup> [10,11]; resistance to *Septoria tritici* on chromosome 4Hc<sup>h</sup> [12]; tolerance to salt on chromosomes 1Hch, 4Hc<sup>h</sup> and 5Hc<sup>h</sup> [13]; fertility restoration on chromosome 6Hc<sup>h</sup> [14]; and carotenoid content on chromosome 7Hc<sup>h</sup> [15]. Wheat—*H. chilense* translocation or recombinant lines have also been generated using both addition and substitution lines of *H. chilense* chromosomes in wheat background [8,11,16–18].

Chromosome 2Hc<sup>h</sup> has the potential to improve seed carotenoid content in wheat. Genetic studies of yellow pigment content (YPC) in *H. chilense* revealed that chromosome 2Hc<sup>h</sup> showed a significant association with YPC [19] and four endosperm carotenoid-related genes have been genetically mapped to chromosome 2Hch, such as geranyl geranyl pyrophosphate synthase (*Ggpps1*) for geranylgeranyl diphosphate synthesis, zeta-carotene desaturase (*Zds*), beta-carotene hydroxylase 3 (*Hyd3*) from the carotenoid biosynthetic pathway and polyphenol oxidase 1 gene (*Ppo1*) implicated in plant tissue enzymatic browning [20].

Molecular markers that are able to distinguish *H. chilense* chromosome 2Hc<sup>h</sup> in wheat background provide a useful tool for selection. The conserved orthologous set (COS) [21] represents an important reservoir of markers that allow comparative studies with wheat and barley and their transference to *H. chilense* is a main goal.

The aims of this work were the following: (a) to obtain wheat—*H. chilense* chromosome 2Hc<sup>h</sup> introgression lines; (b) to characterize the lines obtained by fluorescence *in situ* hybridization (FISH) and chromosome-specific simple sequence repeat (SSR) markers; (c) to transfer COS markers to *H. chilense* and to determine their arm location within 2Hc<sup>h</sup> and (d) to compare the arm location with wheat and barley homoeologous group 2.
