Title: Genetic studies using molecular markers for analysis of complex traits in Phaseolus vulgaris L.
Other Titles: Moleculaire merker studies voor de analyse van complexe eigenschappen in Phaseolus vulgaris L.
Authors: Galeano Mendoza, Carlos
Issue Date: 25-May-2012
Abstract: A major goal of plant genomics is to understand the genetic basis of quantitative traits. This can be achieved by evaluating the association between molecular markers and phenotypic variation by linkage or association mapping approaches. The first phase of this thesis was to develop a reliable genetic map in common bean (Phaseolus vulgaris L.) and to use this map to analyze the inheritance of yield traits under drought and fully irrigated field conditions. This work was conducted using a mapping population consisting of 92 recombinant inbred lines from the Mesoamerican cross DOR364 × BAT477 (DB). The source of drought tolerance used in the cross was the cream-seeded advanced line BAT477. Quantitative trait loci (QTL) were detected for the traits of overall seed yield, yield per day, 100 seed weight, days to flowering and days to maturity for each field environment. The genetic map based on amplified fragment length polymorphism (AFLP) and random amplification of polymorphic DNA (RAPD) was anchored with 60 SSRs markers and had a total map length of 1,087.5 cM across 11 linkage groups. Although this work generated useful QTL results, the linkage map lacked gene-based markers that would allow inferences about the genes involved in the phenotypic responses. Therefore, the second strategy was to develop a set of 418 expressed sequence tag (EST) based amplicons. These markers were evaluated for parental polymorphisms using the single strand conformation polymorphism (SSCP) technique, and 26% of these showed polymorphism between Andean and Mesoamerican genotypes. A total of 118 new marker loci were placed on the DOR364 × G19833 (DG) map, which consisted of 288 markers. Of these, 218 were used for synteny analysis with Glycine max, Medicago, and Lotus. The synteny analysis with soybean showed a mosaic pattern of syntenic blocks with most segments of any one common bean linkage group associated with two soybean chromosomes. The analysis with Medicago truncatula and Lotus japonicus presented fewer syntenic regions consistent with the more distant phylogenetic relationship between the galegoid and phaseoloid legumes.With this new genetic framework and looking for have anchor markers between linkage maps, the third phase was the further saturation of the linkage map of the DB population by evaluating 2,706 additional molecular markers including SSR, single nucleotide polymorphism (SNP), and gene-based markers. On average the polymorphism rate was 7.7% due to the narrow genetic base between the parents. The DB linkage map now consisted of 291 markers with a total map length of 1,788 cM. A consensus map was then constructed using the core mapping populations derived from inter-gene pool crosses: DG and BAT93×JALO EEP558 (BJ). The consensus map consisted of a total of 1,010 markers, with a total map length of 2,041 cM across 11 linkage groups. A synteny analysis was carried out using our highly saturated consensus maps compared with the soybean pseudo-chromosome assembly. A total of 44 syntenic blocks were identified. The linkage group Pv6 presented the most diverse pattern of synteny with seven syntenic blocks, and Pv9 showed the most consistent relations with soybean with just two syntenic blocks. Additionally, a co-linear analysis using common bean transcript map information against soybean coding sequences (CDS) revealed the relationship with 787 soybean genes.Finally, a new set of gene based markers were developed based on an intron-spanning EST-SNP marker approach. In total, 313 new gene-based markers were developed. The markers were evaluated for parental polymorphisms using the SSCP technique and the Sequenom MassARRAY system and 53 loci were placed on the DG linkage map. The new linkage map was merged with the previous consensus map. These new genomic resources were applied to a diversity panel of 93 genotypes. A set of 173 SNP markers was assayed using the Sequenom MassARRAY-platform for association studies. Some significant associations with yield components were identified, and these were consistent with previous findings. Overall, these studies illustrate the power of gene based markers and synteny analysis to develop a framework for analyzing quantitative traits. These results provide a baseline for future efforts in positional cloning of target genes in common bean. Additionally, the consensus map will be a key tool for the eventual assembly of whole genome sequencing in common bean.
Publication status: published
KU Leuven publication type: TH
Appears in Collections:Centre of Microbial and Plant Genetics

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