Beata Myśków

A High Density Consensus Map of Rye (Secale cereale L.) Based on DArT Markers

Background Rye (Secale cereale L.) is an economically important crop, exhibiting unique features such as outstanding resistance to biotic and abiotic stresses and high nutrient use efficiency. This species presents a challenge to geneticists and breeders due to its large genome containing a high proportion of repetitive sequences, self incompatibility, severe inbreeding depression and tissue culture recalcitrance. The genomic resources currently available for rye are underdeveloped in comparison with other crops of similar economic importance. The aim of this study was to create a highly saturated, multilocus linkage map of rye via consensus mapping, based on Diversity Arrays Technology (DArT) markers. Methodology/Principal Findings Recombinant inbred lines (RILs) from 5 populations (564 in total) were genotyped using DArT markers and subjected to linkage analysis using Join Map 4.0 and Multipoint Consensus 2.2 software. A consensus map was constructed using a total of 9703 segregating markers. The average chromosome map length ranged from 199.9 cM (2R) to 251.4 cM (4R) and the average map density was 1.1 cM. The integrated map comprised 4048 loci with the number of markers per chromosome ranging from 454 for 7R to 805 for 4R. In comparison with previously published studies on rye, this represents an eight-fold increase in the number of loci placed on a consensus map and a more than two-fold increase in the number of genetically mapped DArT markers. Conclusions/Significance Through the careful choice of marker type, mapping populations and the use of software packages implementing powerful algorithms for map order optimization, we produced a valuable resource for rye and triticale genomics and breeding, which provides an excellent starting point for more in-depth studies on rye genome organization.

Comparison of RAPD, ISSR and SSR markers in assessing genetic diversity among rye (Secale cereale L.) inbred lines

Comparison of RAPD, ISSR and SSR markers in assessing genetic diversity among rye (Secale cereale L.) inbred lines Forty eight inbred lines of winter rye, of various origin and pedigree, were analysed using 19 RAPD (random amplified polymorphic DNA) primers, 8 ISSR (inter-simple sequence repeats) primers and 13 SSR (simple sequence repeats) primer pairs. On the basis of particular marker types, there were created three separate dendrograms and one combined similarity tree, prepared on account of the whole data. Correlation coefficients for individual technique based on genetic similarity matrices were not significant. By comparing the GS data obtained on the basis of singular methods with collective matrix, it was observed that the highest correlation rate was for ISSR method (r=0.68). The utility of each marker technique was compared by using marker index MI. Diversity detecting index (DDT) was suggested in the paper, which may prove helpful in planning and comparing researches on phenetic relationships.

The application of high-density genetic maps of rye for the detection of QTLs controlling morphological traits

The development of genetic maps is, nowadays, one of the most intensive research activities of plant geneticists. One of the major goals of genome mapping is the localisation of quantitative trait loci (QTLs). This study was aimed at the identification of QTLs controlling morphological traits of rye and comparison of their localisation on genetic maps constructed with the use of genetically different germplasms. For QTL analyses, two high-density consensus maps of two populations (RIL-S and RIL-M) of recombinant inbred lines (RIL) were applied. Plant height (Ph), length of spikes (Sl) and the number of spikelets per spike (Sps) were studied in both populations. Additionally, the number of kernels per spike under isolation (Kps), the weight of kernels per spike (Kw) and thousand kernel weight (Tkw) were assessed in the RIL-M population. Except for Tkw, the majority of the traits were correlated to each other. The non-parametric Kruskal–Wallis (K-W) test and composite interval mapping (CIM) revealed 18/48 and 24/18 regions of rye chromosomes engaged in the determination of Ph, Sl and Sps in the RIL-S and RIL-M populations, respectively. An additional 18/15 QTLs controlling Kps, Kw and Tkw were detected on a map of the RIL-M population. A numerous group of QTLs detected via CIM remained in agreement with the genomic regions found when the K-W test was applied. Frequently, the intervals indicated by CIM were narrower.

A consensus map of chromosome 6R in rye [Secale cereale L.]

Four F2 mapping populations derived from crosses between rye inbred lines DS2×RXL10, 541×Ot1-3, S120×S76 and 544×Ot0-20 were used to develop a consensus map of chromosome 6R. Thirteen marker loci that were polymorphic in more than one mapping population constituted the basis for the alignment of the four maps using the JoinMap v. 3.0 software package. The consensus map consists of 104 molecular marker loci including RFLPs, RAPDs, AFLPs, SSRs, ISSRs, SCARs, STSs and isozymes. The average distance between the marker loci is 1.3 cM, and the total map length is 135.5 cM. This consensus map may be used as a source of molecular markers for the rapid development of new maps of chromosome 6R in any mapping population.

Genetic mapping of the ScHd1 gene in rye and an assessment of its relationship with earliness per se and plant morphology

A fragment of the ScHd1 gene derived from eight inbred lines was sequenced and showed homology to other Hd1 genes from different cereals. Sequences were analysed with respect to the presence of a single-nucleotide polymorphism (SNP) difference. A C-T transition at position 312 of the consensus sequence was found, which distinguished two lines from the remaining six. The deduced amino acid sequence revealed a high identity (93%) to a Hd1-like protein from wheat. The identified mutation allowed the localisation of ScHd1 on a genetic map of rye (6RS). A small, statistically significant linkage between ScHd1 and earliness per se (eps) and some morphological traits was also established. The chromosomal region, including the S76 allele for the ScHd1 gene was linked to earlier heading, elongated spikes, a greater number of spikelets per spike and an increased weight of 1000 kernels.

Phenotypic effect and chromosomal localization of Ddw3, the dominant dwarfing gene in rye (Secale cereale L.)

AbstractDwarfing genes play a major role in development of semi-dwarf cultivars of various cereals, but in rye this type of cultivar is still not common. Several recessive and dominant dwarfing genes in rye have been reported. Among the known dominant dwarfing genes in rye, two are well characterised: Ddw1 from the 5RL chromosome and Ddw2 located on 7R. This study was aimed at characterisation of the K11 source of dominantly inherited dwarfism found in plant materials grown in the Plant Breeding and Acclimatization Institute (Radzików, Poland). Mapping analyses in this study indicate that the dominant dwarf gene under investigation is located on the 1RL chromosome and is independent from previously known genes. The gene was named Ddw3. Phenotypic effect of the Ddw3 was tested on two pairs of near-isogenic lines. Six morphological traits were analysed in two or three growing seasons: plant height, length of the second internode from the base, number of internodes, tillering, spike length, and number of spikelets per spike. No negative or weak influences on yield-related traits were observed. The examination of the dwarf plants’ reaction to seedling treatment by gibberellic acid solution revealed sensitivity of Ddw3 to this growth regulator. This report is the first evidence of the dominant Ddw3 gene in rye and its presence on the 1R chromosome.

Bidirectional selective genotyping approach for the identification of quantitative trait loci controlling earliness per se in winter rye (Secale cereale L.)

The genes controlling earliness of plants include genes responsible for vernalisation (Vrn) and photoperiod (Ppd), and those that are not entirely associated with a response to temperature or light. The last group of loci is known as earliness per se (Eps). Eps genes have been most commonly reported in the scientific literature as quantitative trait loci (QTL). The objective of this study was to use a bidirectional selective genotyping (BSG) method for the identification of loci controlling Eps, assessed at the heading stage in a newly developed rye population of recombinant inbred lines. We identified four linkage groups representing fragments of chromosomes, 1R, 5R, 6R and 7R, and containing, in total, 155 DArT markers significantly associated with earliness in rye. The results are discussed in the context of previously published QTL analyses.

Effective BAC clone anchoring with genotyping-by-sequencing and Diversity Arrays Technology in a large genome cereal rye

Identification of bacterial artificial chromosome (BAC) clones containing specific sequences is a prerequisite for many applications, such as physical map anchoring or gene cloning. Existing BAC library screening strategies are either low-throughput or require a considerable initial input of resources for platform establishment. We describe a high-throughput, reliable, and cost-effective BAC library screening approach deploying genotyping platforms which are independent from the availability of sequence information: a genotyping-by-sequencing (GBS) method DArTSeq and the microarray-based Diversity Arrays Technology (DArT). The performance of these methods was tested in a very large and complex rye genome. The DArTseq approach delivered superior results: a several fold higher efficiency of addressing genetic markers to BAC clones and anchoring of BAC clones to genetic map and also a higher reliability. Considering the sequence independence of the platform, the DArTseq-based library screening can be proposed as an attractive method to speed up genomics research in resource poor species.

Putative candidate genes responsible for leaf rolling in rye ( Secale cereale L.)

BackgroundRolling of leaves (RL) is a phenomenon commonly found in grasses. Morphology of the leaf is an important agronomic trait in field crops especially in rice; therefore, majority of the rice breeders are interested in RL. There are only few studies with respect to RL of wheat and barley; however, the information regarding the genetic base of RL with respect to the shape of leaf in rye is lacking. To the best of our knowledge, this is the first study on the localization of loci controlling RL on high density consensus genetic map of rye.ResultsGenotypic analysis led to the identification of 43 quantitative trait loci (QTLs) for RL, grouped into 28 intervals, which confirms the multigenic base of the trait stated for wheat and rice. Four stable QTLs were located on chromosomes 3R, 5R, and 7R.Co-localization of QTL for RL and for different morphological, biochemical and physiological traits may suggests pleiotropic effects of some QTLs. QTLs for RL were associated with QTLs for such morphological traits as: grain number and weight, spike number per plant, compactness of spike, and plant height. Two QTLs for RL were found to coincide with QTLs for drought tolerance (4R, 7R), two with QTLs for heading earliness (2R, 7R), one with α-amylase activity QTL (7R) and three for pre-harvest sprouting QTL (1R, 4R, 7R).The set of molecular markers strongly linked to RL was selected, and the putative candidate genes controlling the process of RL were identified. Twelve QTLs are considered as linked to candidate genes on the base of DArT sequences alignment, which is a new information for rye.ConclusionsOur results expand the knowledge about the network of QTLs for different morphological, biochemical and physiological traits and can be a starting point to studies on particular genes controlling RL and other important agronomic traits (yield, earliness, pre-harvest sprouting, reaction to water deficit) and to appoint markers useful in marker assisted selection (MAS). A better knowledge of the rye genome and genes could both facilitate rye improvement itself and increase the efficiency of utilizing rye genes in wheat breeding.