A High-Density Linkage Map of the Forage Grass Eragrostis curvula and Localization of the Diplospory Locus

Abstract

Eragrostis curvula (Schrad.) Nees (weeping lovegrass) is an apomictic species native to Southern Africa that is used as forage grass in semiarid regions of Argentina. Apomixis is a mechanism for clonal propagation through seeds that involves the avoidance of meiosis to generate an unreduced embryo sac (apomeiosis), parthenogenesis, and viable endosperm formation in a fertilization-dependent or -independent manner. Here, we constructed the first saturated linkage map of tetraploid E. curvula using both traditional (AFLP and SSR) and high-throughput molecular markers (GBS-SNP) and identified the locus controlling diplospory. We also identified putative regulatory regions affecting the expressivity of this trait and syntenic relationships with genomes of other grass species. We obtained a tetraploid mapping population from a cross between a full sexual genotype (OTA-S) with a facultative apomictic individual of cv. Don Walter. Phenotypic characterization of F 1 hybrids by cytoembryological analysis yielded a 1:1 ratio of apomictic vs. sexual plants (34:27, X 2 = 0.37), which agrees with the model of inheritance of a single dominant genetic factor. The final number of markers was 1,114 for OTA-S and 2,019 for Don Walter. These markers were distributed into 40 linkage groups per parental genotype, which is consistent with the number of E. curvula chromosomes (containing 2 to 123 markers per linkage group). The total length of the OTA-S map was 1,335 cM, with an average marker density of 1.22 cM per marker. The Don Walter map was 1,976.2 cM, with an average marker density of 0.98 cM/marker. The locus responsible for diplospory was mapped on Don Walter linkage group 3, with other 65 markers. QTL analyses of the expressivity of diplospory in the F 1 hybrids revealed the presence of two main QTLs, located 3.27 and 15 cM from the diplospory locus. Both QTLs explained 28.6% of phenotypic variation. Syntenic analysis allowed us to establish the groups of homologs/homeologs for each linkage map. The genetic linkage map reported in this study, the first such map for E. curvula , is the most saturated map for the genus Eragrostis and one of the most saturated maps for a polyploid forage grass species.

Genes Modulating the Increase in Sexuality in the Facultative Diplosporous Grass Eragrostis curvula under Water Stress Conditions

“…Although an E. curvula genome assembly is available [ 55 ] and we could have used it as a reference genome, for this study we decided to de novo assemble the reads since the sequenced genome belongs to a sexual diploid genotype and the sequences corresponding to the region that determine apomixis might not be present, as was observed by Zappacosta et al [ 54 ] using molecular markers linked to the trait. The assembly quality analysis, performed with Trinity stats, indicated an N50 of 1553 bp, an average contig length of 919.09 bp and a GC content of 46.38%.…”

“…Finally, both treatments were conducted with a facultative apomictic cultivar, hence we expected to find mostly apomictic/sexual regulators, not apomictic triggers because they must be present in plants from both treatments. On the other hand, we are working with the knowledge that apomixis in E. curvula is determined by a genomic region [ 54 ] with an epigenetic component modulating the traits [ 40 , 52 ], and apomixis and sexuality co-exist in facultative plants [ 54 ], allowing us to hypothesize that sexuality is being repressed at different levels in apomictic plants. This repression is deregulated by different endogenous [ 56 ] and exogenous stresses [ 40 ], probably mediated by transcriptional and post transcriptional regulatory mechanisms.…”

“…Eragrostis curvula (weeping lovegrass), an African grass with cytotypes of different ploidy levels (e.g., 2x–8x) and displaying obligate and facultative apomixis and sexual reproduction [ 46 ], has become a model for the analysis of apomixis mechanisms, due to its particular diplosporous development (meiotic diplospory maintaining the same embryo: endosperm ploidy ratio as in sexual seeds). In recent years, the reproductive mode of this grass was studied extensively, providing information about the cytoembryological aspects of its apomictic–sexual development [ 47 ], differentially expressed (DE) transcripts [ 48 , 49 , 50 , 51 ], epigenetic aspects of apomictic regulation [ 52 , 53 ], mapping of the apomixis locus [ 54 ] and a high quality genome assembly [ 55 ]. Our group also demonstrated that under different internal and external stressful situations, including a change in ploidy, water stress, in vitro culture and intraspecific hybridization, the number of sexual embryo sacs increased in facultative apomictic plants of this grass [ 40 , 56 ].…”

Controlling Apomixis: Shared Features and Distinct Characteristics of Gene Regulation

“…Direct identification of the genes and genomic elements comprised on these apomixis linked loci however has proven difficult. This is because they are commonly recombination-suppressed and flanked by repetitive regions, interfering with sequencing approaches and map-based cloning [10,[35][36][37][38][39][40][41]. In several species studied, apomixis linked loci represent chromosomal regions that largely diverged from corresponding sexual loci.…”

Differential Methylation Patterns in Apomictic vs. Sexual Genotypes of the Diplosporous Grass Eragrostis curvula

“…This is due to its particular type of embryo sac development (Eragrostis-type) [ 30 ]. The reproductive mode of this grass was widely studied using different approaches like cyto-embryology [ 31 ], differential gene expression [ 32 , 33 , 34 ], mapping of the apomixis locus [ 35 ], and genome assembly [ 36 ]. Moreover, it was shown that internal and external stressful situations, like water stress, in vitro culture and intraspecific hybridization increase the percentage of sexual embryo sacs in facultative apomictic plants, evidencing the existence of some epigenetic control over the process [ 37 , 38 ].…”