**5. Conclusions**

We used in situ hybridization and genotyping to characterize genetic stocks harboring chromosome 2Hc<sup>h</sup> from *H. chilense* in a wheat background. As far as we know, no previous translocation lines involving chromosome 2Hchintrogressed into wheat have been described. The cytogenetic stocks developed here may constitute an important resource for studying the effect of chromosome 2Hc<sup>h</sup> on wheat grain color. In addition, these cytogenetic stocks allowed the localization of a set of conserved orthologous set (COS) markers to specific arms in chromosome 2Hch.The genomic position of orthologous unigene EST-contigs used for the COS marker design revealed a good macrosyntenic relationship between *H. chilense* chromosome 2Hc<sup>h</sup> and wheat homoeologous group 2. The new wheat—*H. chilense* recombinations are useful for genetic studies and might also serve as a bridge for transferring genes associated with yellow pigment into a wheat background.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2073-4395/9/9/493/s1, File S1: COS markers used in this work together with their primer sequences, File S2: Source of the COS markers assigned to the *H. chilense* chromosome 2Hch, File S3: Results of BLASTn search for COS markers assigned to *H. chilense* chromosome 2Hc<sup>h</sup> in the reference sequences of hexaploid wheat group 2 chromosomes (www.wheatgenome.org/) and the start positions (bp) of the marker-specific ESTs.

**Author Contributions:** A.C. conceived and designed the study; C.P. performed the experiments; A.C. analysed the data and wrote the paper: all authors have read and approved the final manuscript.

**Funding:** This research was supported by grants AGL2014-53195-R and RTI2018-093367-B-I00 from the Spanish State Research Agency (Ministry of Science, Innovation and Universities), co-financed by the European Regional Development Fund (FEDER) from the European Union.

**Conflicts of Interest:** The authors declare no conflict of interest.
