S-SAP molecular marker-based detection of interspecific variability among grape cultivars

Genetska unutarsortna varijabilnost temelji se na pretpostavci da tijekom vremena dolazi do pojave spontanih mutacija u genomu sorte koje zbog vegetativnog načina razmnožavanja ostaju fiksirane. Svaka informacija o unutarsortnoj varijabilnosti može biti od velike važnosti prije odluke o tome koje trsove zadržati nakon provjere zdravstvenog statusa te može pružiti uvid u homogenost populacije i potencijal klonske selekcije. U ovom istraživanju korišten je sustav markera temeljen na retrotranspozonima – modificirana S-SAP metoda koja je u prijašnjim istraživanjima pokazala potencijal za razlikovanje klonova. Korišten biljni materijal vinove loze sastojao se od 22 primke dviju različitih sorti: Zlatarice blatske bijele i Glavinuše. S-SAP metoda provedena je sa 6 različitih kombinacija početnica. Ovom metodom uspješno su razlikovane obje sorte, a detektirane su i razlike među pojedinim jedinkama. Stupanj polimorfizma, iako analize nisu rađene na certificiranom klonskom materijalu, u skladu je sa sličnim istraživanjima rađenim na certificiranom materijalu. Početnica F100 je detektirala najviše razlika i može se preporučiti u daljnjim istraživanjima. Dobiveni rezultati pružaju smjernice prilikom daljnje propagacije i revitalizacije.Intra-varietal variability is based on the assumption that during the time spontaneous mutations occur in the varieties genome and stay fixed because of vegetative propagation. Any kind of information about intra-varietal variability could be of importance before making a decision which vines to keep after verification of their health status and could provide an insight into homogeneity of the population and into the potential of the clonal selection. In this study a retrotransposon based marker system was used – a modified SSAP method which, in previous studies, showed the potential for differentiating clones. Collected grapevine plant material consisted of 22 accessions representing two Croatian varieties: Zlatarica blatska bijela and Glavinuša. S-SAP method was performed using 6 different primer combinations. This method successfully differentiated both varieties and also detected some clonal differences among tested accessions. Although the analyses were not done on certified clonal material, level of polymorphism was similar to those in similar studies using certified clones. The use of universal retrotransposon primer F100 resulted in most observed polymorphism. Obtained results provide guidelines for further propagation and revitalization

Characteristaion of the initial generations of recombinant inbred lines in perennial ryegrass (lolium perenne L.) using molecular markers and cytogenetics

Doctoral ThesisIn this study three early generations of a recombinant inbred line (RIL) breeding programme have been characterised by cytogenetic methods, metabolite profiling and a biomass quantitative trait loci (QTL) study. In situ hybridization was used to analyse recombination and the structure of the parental and the F1 genomes. A metabolite profiling study via GC/MS was conducted to characterise the metabolite activity of the parental inbred lines and the F1 hybrid. Total metabolites were compared across a growth season in a replicated field design. A genetic map was constructed with 75 nuclear DNA markers in the F2 generation, which consisted of 360 individuals. 63% of the markers in the F2 population did not fit the expected Mendelian ratios and showed significant (p<0.05) segregation distortion. Fresh weight, dry weight, and dry matter were analysed with an alpha lattice design throughout the greenhouse and field experiments. Additionally, leaf width was recorded in the greenhouse experiment. Heritablities ranged for the biomass related traits between 78 and 95%. This study provides an insight into the recombination of the parental and the F1 genotypes. Different levels of metabolite activities could be found among the two parental inbred lines across three harvest dates. The QTL study indicates the position of biomass QTL related traits. Major QTL with log of odds scores >3 were identified on linkage groups 2, 3, 4, 5 and 7. About 30% of overall variation could be explained. Increased biomass yield is still one of the most important traits in any Lolium perenne breeding programme. The present QTL study can be used for fine mapping of biomass yield related traits in L. perenne. In the long term biomass yield can be eventually monitored and predicted with marker assisted selection for some of the QTL identified in the present study.Teagasc Walsh Fellowship programme; National Development Plan Irelan

Characterisation of the initial generations of recombinant inbred lines in perennial ryegrass (Lolium perenne L.) using molecular markers and cytogenetics

Teagasc for the Walsh Fellowship and the National Development Plan for financial contributionsEThOS - Electronic Theses Online ServiceTeagascGBUnited Kingdo

An Immortalized Genetic Mapping Population for Perennial Ryegrass: A Resource for Phenotyping and Complex Trait Mapping.

To address the lack of a truly portable, universal reference mapping population for perennial ryegrass, we have been developing a recombinant inbred line (RIL) mapping population of perennial ryegrass derived via single seed descent from a well-characterized F2 mapping population based on genetically distinct inbred parents in which the natural self-incompatibility (SI) system of perennial ryegrass has been overcome. We examined whether it is possible to create a genotyping by sequencing (GBS) based genetic linkage map in a small population of the F6 generation of this population. We used 41 F6 genotypes for GBS with PstI/MspI-based libraries. We successfully developed a genetic linkage map comprising 6074 SNP markers, placing a further 22080 presence and absence variation (PAV) markers on the map. We examined the resulting genetic map for general and RIL specific features. Overall segregation distortion levels were similar to those experienced in the F2 generation, but segregation distortion was reduced on linkage group 6 and increased on linkage group 7. Residual heterozygosity in the F6 generation was observed at a level of 5.4%. There was a high proportion of chromosomes (30%) exhibiting the intact haplotype of the original inbred parents of the F1 genotype from which the population is derived, pointing to a tendency for chromosomes to assort without recombining. This could affect the applicability of these lines and might make them more suitable for situations where repressed recombination is an advantage. Inter- and intra-chromosomal linkage disequilibrium (LD) analysis suggested that the map order was robust. We conclude that this RIL population, and subsequent F7 and F8 generations will be useful for genetic analysis and phenotyping of agronomic and biological important traits in perennial ryegrass

Data_Sheet_3_An Immortalized Genetic Mapping Population for Perennial Ryegrass: A Resource for Phenotyping and Complex Trait Mapping.docx

<p>To address the lack of a truly portable, universal reference mapping population for perennial ryegrass, we have been developing a recombinant inbred line (RIL) mapping population of perennial ryegrass derived via single seed descent from a well-characterized F2 mapping population based on genetically distinct inbred parents in which the natural self-incompatibility (SI) system of perennial ryegrass has been overcome. We examined whether it is possible to create a genotyping by sequencing (GBS) based genetic linkage map in a small population of the F6 generation of this population. We used 41 F6 genotypes for GBS with PstI/MspI-based libraries. We successfully developed a genetic linkage map comprising 6074 SNP markers, placing a further 22080 presence and absence variation (PAV) markers on the map. We examined the resulting genetic map for general and RIL specific features. Overall segregation distortion levels were similar to those experienced in the F2 generation, but segregation distortion was reduced on linkage group 6 and increased on linkage group 7. Residual heterozygosity in the F6 generation was observed at a level of 5.4%. There was a high proportion of chromosomes (30%) exhibiting the intact haplotype of the original inbred parents of the F1 genotype from which the population is derived, pointing to a tendency for chromosomes to assort without recombining. This could affect the applicability of these lines and might make them more suitable for situations where repressed recombination is an advantage. Inter- and intra-chromosomal linkage disequilibrium (LD) analysis suggested that the map order was robust. We conclude that this RIL population, and subsequent F7 and F8 generations will be useful for genetic analysis and phenotyping of agronomic and biological important traits in perennial ryegrass.</p