A high carotenoid content is important for the production of pasta from durum wheat (Triticum durum Desf.) and yellow alkaline noodle from common wheat (T. aestivum L.). Carotenoid esters are more stable than free carotenoid during storage and processing, and thus they allow a higher retention through the food chain. Chromosome 7D carries gene(s) for lutein esterification. The aim of this study was the physical mapping of the gene(s) for lutein esterification on chromosome 7D and the identification of candidate genes for this trait. We developed crosses between a set of deletion lines for chromosome 7D in Chinese Spring (CS) background and the CS–Hordeum chilense substitution line CS(7D)7Hch. The F2 progeny derived from the deletion line 7DS4 produced a lower amount of lutein esters, which indicates that the main gene for lutein esterification is in the region of chromosome 7D lacking in 7DS4. Other gene(s) are contributing to lutein esterification because small amounts of lutein esters are produced in 7DS4. Genotyping by DArTSeq revealed that 7DS4 lacks a 127.7 Mb region of 7DS. A set of 10 candidate genes for lutein esterification was identified by using the wheat reference genome sequence along with the Wheat Expression Browser. This region contains the Lute locus previously identified in a different genetic background. Four genes with acyltransferase or GDSL esterase/lipase activity were identified in the vicinity of Lute. Our results indicate that the gene TraesCS7D01G094000 is a likely candidate for Lute but the gene TraesCS7D01G093200 cannot be ruled out. The candidate genes reported in this work are worthy for further investigation., This research was funded by Grants AGL2014-53195-R from Ministerio de Economía y Competitividad and AGL2017-85368-P from Ministerio de Ciencia, Innovación y Universidades. Both projects are cofounded by FEDER. DHM is member of CaRed Network funded by Ministerio de Economía y Competitividad (BIO2015-71703-REDT and BIO2017-90877-REDT) and participates in EUROCAROTEN COST Action (CA15136). CMA and SGA are members of FiRCMe Network, funded by Ministerio de Economía y Competitividad (AGL2016-81855-REDT)., Peer reviewed

3 Figuras, Tritordeums show a significant proportion of lutein esters which increases carotenoid stability and retention throughout the food chain. Esterification is a common means of carotenoid sequestration. A putative association between lutein esters formation acting as a metabolic sink during early stages of grain development and the high carotenoid content of tritordeums is analyzed in this work.

Compared to wheat, tritordeums accumulated significantly higher lutein contents from 20 days post anthesis (dpa) but lutein esters were not detected until 36 dpa. Thus esterification is not acting as a metabolific sink before 36 dpa. The presence of lutein esters at late stages of grain development may have a complementary role in carotenoid accumulation by reducing and/or counteracting their catabolism. The differences for lutein esterification among tritordeums suggest the existence of diversity for xanthophyll acyl transferases that could be exploited to increase lutein retention in this cereal and through the food chain., This research was funded by Grants AGL2014-53195-R from Ministerio de Economía y Competitividad and AGL2017-85368-P from Ministerio de Ciencia, Innovación y Universidades. Both projects are cofounded by FEDER. DHM is member of CaRed Network funded by Ministerio de Economía y Competitividad (BIO2015-71703-REDT and BIO2017-90877-REDT) and participates in EUROCAROTEN COST Action (CA15136). SGA is member of FiRCMe Network, funded by Ministerio de Economía, Industria y Competitividad (AGL2016-81855-REDT). M.G.M. was recipient of a fellowship from Ministerio de Economía y Competitividad (BES-2012-055961)., Peer reviewed

Carotenoids are essential in human diet, so that the development of programs toward carotenoid enhancement has been promoted in several crops. The cereal tritordeum, the amphiploid derived from the cross between Hordeum chilense Roem. et Schulz. and durum wheat has a remarkable carotenoid content in the endosperm. Besides, a high proportion of these carotenoids are esterified with fatty acids. The identification of the gene(s) responsible for xanthophyll esterification would be useful for breeding as esterified carotenoids show an increased ability to accumulate within plant cells and have a higher stability during post-harvest storage. In this work, we analyzed five genes identified as candidates for coding the xanthophyll acyltransferase (XAT) enzyme responsible for lutein esterification in H. chilense genome. All these genes were expressed during grain development in tritordeum, but only HORCH7HG021460 was highly upregulated. Sequence analysis of HORCH7HG021460 revealed a G-to-T transversion, causing a Glycine to Cysteine substitution in the protein of H290 (the only accession not producing quantifiable amounts of lutein esters, hereinafter referred as zero-ester) of H. chilense compared to the esterifying genotypes. An allele-specific marker was designed for the SNP detection in the H. chilense diversity panel. From the 93 accessions, only H290 showed the T allele and the zero-ester phenotype. Furthermore, HORCH7HG021460 is the orthologue of XAT-7D, which encodes a XAT enzyme responsible for carotenoid esterification in wheat. Thus, HORCH7HG021460 (XAT-7Hch) is a strong candidate for lutein esterification in H. chilense and tritordeum, suggesting a common mechanism of carotenoid esterification in Triticeae species. The transference of XAT-7Hch to wheat may be useful for the enhancement of lutein esters in biofortification programs., This research was funded by grant AGL2017-85368-P from Ministerio de Ciencia, Innovación y Universidades, and co-funded by FEDER. MR-R was recipient of a fellowship from Ministerio de Ciencia, Innovación y Universidades (PRE2018-084037). DH-M is member of CaRed Network, funded by Ministerio de Economía y Competitividad (BIO2017-90877-REDT) and participated in EUROCAROTEN COSTAction (CA15136). SA is member of FiRCMe Network, funded by Ministerio de Economía, Industria y Competitividad (AGL2016-81855-REDT)., Peer reviewed

10 Páginas.-- 4 Figuras.-- 1 Tabla, Carotenoids are essential in the human diet for their important functions in health. Besides, they are responsible for the yellow pigments desirable for industrial quality in durum wheat. The remarkable carotenoid content of durum wheat endosperm is mostly due to lutein. However, lutein esters have not been previously detected in durum wheat as in other cereals such as common wheat, tritordeum or Hordeum chilense. Esterification increases carotenoid stability and allows greater retention and accumulation through the food chain. Therefore, carotenoid esterification is revealed as a new key target in breeding. We characterized the carotenoid profile of 156 accessions of the Spanish durum wheat collection, searching for landraces with esterification ability. Interestingly, four accessions produced lutein monoesters and diesters. Also, traces of lutein monoesters were detected in eleven accessions. The identification of the first durum wheat accessions with esterification ability reported herein is a remarkable advance for carotenoid biofortification. Furthermore, variation for the relative content of zeaxanthin, α-carotene and β-carotene was also observed. This diversity for the β,ε and β,β branches of the carotenogenic pathway also represents a new opportunity for breeding for specific carotenoids in biofortification programs., This research was funded by Ministerio de Ciencia, Innovación y Universidades, grant number AGL2017-85368-P and by Instituto de Investigación y Formación Agraria y Pesquera (IFAPA), Junta de Andalucía, grant number PP.AVA-AVA2019.020. Both grants are cofounded by FEDER. D.H.-M. is member of CaRed Network funded by Ministerio de Economía y Competitividad (BIO2017-90877-REDT) and participates in EUROCAROTEN COST Action (CA15136). S.G.A. is member of FiRCMe Network, funded by Ministerio de Economía, Industria y Competitividad (AGL2016-81855-REDT). M.D.R.-R. was recipient of a fellowship from Ministerio de Ciencia, Innovación y Universidades (PRE2018-084037).

This article belongs to the Special Issue Pre-breeding towards the Effective Utilization of Plant Genetic Resources., Hexaploid tritordeum is the amphiploid derived from the cross between the wild barley Hordeum chilense and durum wheat. This paper reviews the main advances and achievements in the last two decades that led to the successful development of tritordeum as a new crop. In particular, we summarize the progress in breeding for agronomic performance, including the potential of tritordeum as a genetic bridge for wheat breeding; the impact of molecular markers in genetic studies and breeding; and the progress in quality and development of innovative food products. The success of tritordeum as a crop shows the importance of the effective utilization of plant genetic resources for the development of new innovative products for agriculture and industry. Considering that wild plant genetic resources have made possible the development of this new crop, the huge potential of more accessible resources, such as landraces conserved in gene banks, goes beyond being sources of resistance to biotic and abiotic stresses. In addition, the positive result of tritordeum also shows the importance of adequate commercialization strategies and demonstrative experiences aimed to integrate the whole food chain, from producers to end-point sellers, in order to develop new products for consumers., This research was funded by Ministerio de Ciencia, Innovación y Universidades, grant number AGL2017-85368-P and cofounded by FEDER.

15 Páginas.-- 5 Figuras.-- 1 Tabla, Yellow pigment content is one of the main traits considered for grain quality in durum wheat (Triticum turgidum L.). The yellow color is mostly determined by carotenoid pigments, lutein being the most abundant in wheat endosperm, although zeaxanthin, α-carotene and β-carotene are present in minor quantities. Due to the importance of carotenoids in human health and grain quality, modifying the carotenoid content and profile has been a classic target. Landraces are then a potential source for the variability needed for wheat breeding. In this work, 158 accessions of the Spanish durum wheat collection were characterized for carotenoid content and profile and genotyped using the DArTSeq platform for association analysis. A total of 28 marker-trait associations were identified and their co-location with previously described QTLs and candidate genes was studied. The results obtained confirm the importance of the widely described QTL in 7B and validate the QTL regions recently identified by haplotype analysis for the semolina pigment. Additionally, copies of the Zds and Psy genes on chromosomes 7B and 5B, respectively, may have a putative role in determining zeaxanthin content. Finally, genes for the methylerythritol 4-phosphate (MEP) and isopentenyl diphosphate (IPPI) carotenoid precursor pathways were revealed as additional sources of untapped variation for carotenoid improvement., This research was funded by project AGL2017-85368-P funded by MCIN/AEI/10.13039/501100011033/, by ERDF “ERDF A way of making Europe” and by the Instituto de Investigación y Formación Agraria y Pesquera (IFAPA), Junta de Andalucía, grant number AVA-AVA2019.020 cofounded at 80% by ERDF. M.D.R.-R. was supported by PRE2018-084037 funded by MCIN/AEI/10.13039/501100011033 and ESF “ESF investing in your future”., Peer reviewed

11 Páginas.-- 3 Figuras.-- 2 Tablas, Bread wheat has traditionally been selected for whitish derived flours. As a consequence, the current varieties carry carotenogenic alleles associated with low grain carotenoid. In contrast, high grain yellow pigment content (YPC) has been a major target in durum wheat programs since yellow colour is an important aesthetic factor for pasta production. Phytoene synthase 1 (Psy1) genes have an important role in the determination of the carotenoid content in wheat. In this work, we have transferred the genes Psy1-A1 and Psy1-B1 from durum to bread wheat by inter-specific hybridization in order to evaluate the combined effect of these genes for the improvement of grain carotenoid content, as well as the development of carotenoid-enriched bread wheat lines. Inter-specific breeding coupled with a MAS approach based on Psy1-A1 and Psy1-B1 alleles has allowed the development of bread wheat pre-breeding lines with enhanced grain carotenoid content (16-23% mean). These biofortified lines have the potential to become new varieties or to be used as recurrent parents in bread wheat breeding programs., This research was funded by projects AGL2017-85368-P and PID2021-122152NB-I00 funded by MCIN/AEI/10.13039/501100011033/ and by ERDF “ERDF A way of making Europe”. M.D.R.-R. was supported by PRE2018-084037 funded by MCIN/AEI/10.13039/501100011033 and ESF “ESF investing in your future”., Peer reviewed