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Rye (Secale cereale L.) is an exceptionally climate-resilient cereal crop, used extensively to produce improved wheat varieties via introgressive hybridization and possessing the entire repertoire of genes necessary to enable hybrid breeding. Rye is allogamous and only recently domesticated, thus giving cultivated ryes access to a diverse and exploitable wild gene pool. To further enhance the agronomic potential of rye, we produced a chromosome-scale annotated assembly of the 7.9-gigabase rye genome and extensively validated its quality by using a suite of molecular genetic resources. We demonstrate applications of this resource with a broad range of investigations. We present findings on cultivated rye's incomplete genetic isolation from wild relatives, mechanisms of genome structural evolution, pathogen resistance, low-temperature tolerance, fertility control systems for hybrid breeding and the yield benefits of rye-wheat introgressions.

The gibberellin (GA)-sensitive dwarfing gene Ddw1 provides an opportunity to genetically reduce plant height in rye. Genetic analysis in a population of recombinant inbred lines confirmed a monogenetic dominant inheritance of Ddw1 . Significant phenotypic differences in PH between homo- and heterozygotic genotypes indicate an incomplete dominance of Ddw1 . De novo transcriptome sequencing of Ddw1 mutant as well as tall genotypes resulted in 113,547 contigs with an average length of 318 bp covering 36.18 Mbp rye DNA. A hierarchical cluster analysis based on individual groups of rye homologs of functionally characterized rice genes controlling morphological or physiological traits including plant height, flowering time, and source activity identified the gene expression profile of stems at the begin of heading to most comprehensively mirror effects of Ddw1 . Genome-wide expression profiling identified 186 transcripts differentially expressed between semi-dwarf and tall genotypes in stems. In total, 29 novel markers have been established and mapped to a 27.2 cM segment in the distal part of the long arm of chromosome 5R. Ddw1 could be mapped within a 0.4 cM interval co-segregating with a marker representing the C20-GA2-oxidase gene ScGA2ox12 , that is up-regulated in stems of Ddw1 genotypes. The increased expression of ScGA2ox12 observed in semi-dwarf rye as well as structural alterations in transcript sequences associated with the ScGA2ox12 gene implicate, that Ddw1 is a dominant gain-of-function mutant. Integration of the target interval in the wheat reference genome sequence indicated perfect micro-colinearity between the Ddw1 locus and a 831 kb segment on chromosome 5A, which resides inside of a 11.21 Mb interval carrying the GA-sensitive dwarfing gene Rht12 in wheat. The potential of Ddw1 as a breeder's option to improve lodging tolerance in rye is discussed.

The post-zygotic reproductive isolation (RI) in plants is frequently based on the negative interaction of the parental genes involved in plant development. Of special interest is the study of such types of interactions in crop plants, because of the importance of distant hybridization in plant breeding. This study is devoted to map rye genes that are incompatible with wheat, determining the development of the shoot apical meristem in wheat⁻rye hybrids. Linkage analysis of microsatellite loci, as well as genes of embryo lethality ( Eml-R1 ) and hybrid dwarfness ( Hdw-R1 ) was carried out in hybrids of Chinese Spring wheat with recombinant inbred lines as well as interline rye hybrids. Eml-R1 and Hdw-R1 could be mapped proximal and distal of two closely linked EST-SSR markers, Xgrm902 and Xgrm959 , on rye chromosome 6R. Both rye genes are located on a segment of chromosome 6R that contains a breakpoint of evolutionary translocation between the ancestral chromosomes of homeologous groups 6 and 3. The obtained results are discussed in relation to genes interacting in developmental pathways as a class of causal genes of RI.

The color of grain in cereals is determined mainly by anthocyanin pigments. A large level of genetic diversity for anthocyanin content and composition in the grain of different species was observed. In rye, recessive mutations in six genes (vi1...vi6) lead to the absence of anthocyanins in all parts of the plant. Moreover, dominant genes of anthocyanin synthesis in aleurone (gene C) and pericarp (gene Vs) also affect the color of the grain. Reverse phase high-performance liquid chromatography and mass spectrometry were used to study anthocyanins in 24 rye samples. A lack of anthocyanins in the lines with yellow and brown grain was determined. Delphinidin rutinoside and cyanidin rutinoside were found in the green-seeded lines. Six samples with violet grains significantly varied in terms of anthocyanin composition and content. However, the main aglycone was cyanidin or peonidin in all of them. Monosaccharide glucose and disaccharide rutinose served as the glycoside units. Violet-seeded accession forms differ in the ratio of the main anthocyanins and the range of their acylated derivatives. The acyl groups were presented mainly by radicals of malonic and sinapic acids. For the colored forms, a profile of the revealed anthocyanins with the indication of their contents was given. The obtained results are discussed in connection to similar data in rice, barley, and wheat, which will provide a perspective for future investigations.

Six nonallelic genes have been discovered in rye, the recessive mutations of which lead to a lack of anthocyanin. Crosses with these mutants showed that 13 new anthocyaninless lines carry mutations in the gene vi1, whereas vi2/6 mutations were identified only in single cases. Inheritance of the vi1/6 mutations in the progeny of hybrids with the wild type (Line 7, L7) corresponds to a monohybrid segregation. Segregation for three of these mutations (vi2, vi4, and vi5) in hybrids with line 2 is characterized by anthocyaninless deficiency in plants. We discuss the reasons for such deviations and the previously published data for the identification of the six anthocyanin pigmentation genes in the rye using trisomic analysis.

The wheat and rye spike normally bears one spikelet per rachis node, and the appearance of supernumerary spikelets is rare. The loci responsible for the 'multirow spike' or MRS trait in wheat, and the 'monstrosum spike' trait in rye were mapped by genotyping F(2) populations with microsatellite markers. Both MRS and the 'monstrosum' trait are under the control of a recessive allele at a single locus. The Mrs1 locus is located on chromosome 2DS, co-segregating with the microsatellite locus Xwmc453. The placement of flanking microsatellite loci into chromosome deletion bin 2DS-5 (FL 0.47-1.0) delimited the physical location of Mrs1 to the distal half of chromosome arm 2DS, within the gene rich region 2S0.8. The Mo1 locus maps about 10 cM from the centromere on chromosome arm 2RS. The similar effect on phenotype of mo1 and mrs1, together with their presence in regions of conserved synteny, suggest that they may well be members of an orthologous set of Triticeae genes governing spike branching. The practical importance of the MRS spike is that it produces more spikelets per spike, and thereby enhances the sink capacity of wheat, which is believed to limit the yield potential of the crop.

Segregation for self-fertility has been studied in progenies from the crosses of self-sterile (SS) plants with interline hybrids obtained by a diallel scheme of pollinations between seven self-fertile (SF) lines (nos. 2-8) and with F1 (SS plant x SF line) hybrids. All the offspring families from the SS plant x F1 (SS plant x SF line) crosses demonstrated a 1SF∶1SS segregation. The crosses of SS plants with some interline hybrids gave only self-fertile plants, whereas the crosses with other interline hybrids gave a segregation of 3SF:1SS expected in the case of digenic segregation. The data obtained permitted us to identify three different S loci (S1, S2, S5) and to estimate the genotypes of self-fertile lines for their Sf alleles: lines 5, 6, 7 and 8 are S1f/S1f S2n/S2n S5m/S5m, line 4 is S1n/S1n S2f/S2f S5m/S5m, and lines 2 and 3 are S1n/S1n S2m/S2m S5f/S5f(Sn, Sm designate active alleles of the incompatibility genes). The identification of the particular S gene which is presented by the Sf allele in each line has been made on the basis of our data concerning the linkage of the Sf mutation with isozyme markers of particular rye chromosomes, which is reported in an accompanying paper.

The segregation of several isozyme marker genes has been studied in F2 inbred families from hybrids between self-sterile and five self-fertile inbred lines (nos. 2, 3, 4, 5, and 8) as well as from interline hybrids. Self-pollination of F1 hybrids between self-sterile forms and lines 5 and 8 gave an F2 segregation ratio of 1 heterozygote:1 homozygote for the gene Prx7 (chromosome 1R) against the allele from the line. This is interpreted as a result of tight linkage of the Prx7 gene with the S1 gene in chromosome 1R (recombination at a level of 0-1%). The self-pollination of such hybrids with lines 2,3 and 4 gave normal segregation for the Prx7 gene (1:2:1). This means that these lines carry a self-fertility allele which is not on chromosome 1R. Interline hybrids 5×2, 5×3 and 5×4 had self-fertility alleles for the two S genes and in inbred F2 progenies gave the expected deviating segregation for the Prx7 gene in a ratio of 2:3:1. The segregation of interline hybrid 5×8 was normal, 1:2:1, as expected. Highly-deviating segregation in an inbred F2 family of a hybrid with line 5 has also been obtained for another gene from chromosome 1R - Pgi2 (recombination with the S1 locus of 16.7%). By using the same method it has been estimated that line 4 has a self-fertility allele of the S2 locus from chromosome 2R and that the genes β-Glu and Est4/11 are linked with it (recombination 16.7% and 17.5-20% respectively). Lines 2 and 3 have a self-fertility allele of the S5 locus from chromosome 5R which is linked with the Est5-7 gene complex (recombination at a level of 28.8-36.0%).