Association mapping is a method for detection of gene effects based on linkage disequilibrium (LD) that complements QTL analysis in the development of tools for molecular plant breeding. In this study, association mapping was performed on a selected sample of 95 cultivars of soft winter wheat. Population structure was estimated on the basis of 36 unlinked simple-sequence repeat (SSR) markers. The extent of LD was estimated on chromosomes 2D and part of 5A, relative to the LD observed among unlinked markers. Consistent LD on chromosome 2D was <1 cM, whereas in the centromeric region of 5A, LD extended for ∼5 cM. Association of 62 SSR loci on chromosomes 2D, 5A, and 5B with kernel morphology and milling quality was analyzed through a mixed-effects model, where subpopulation was considered as a random factor and the marker tested was considered as a fixed factor. Permutations were used to adjust the threshold of significance for multiple testing within chromosomes. In agreement with previous QTL analysis, significant markers for kernel size were detected on the three chromosomes tested, and alleles potentially useful for selection were identified. Our results demonstrated that association mapping could complement and enhance previous QTL information for marker-assisted selection.

https://www.genetics.org/content/genetics/172/2/1165.full.pdf

Association Mapping of Kernel Size and Milling Quality in Wheat (Triticum aestivum L.) Cultivars

To make advances in rice breeding it is important to understand the relatedness and ancestry of introduced rice accessions, and identify SSR markers associated with agronomically important phenotypic traits, for example yield. Ninety-two rice germplasm accessions recently introduced from seven geographic regions of Africa, Asia, and Latin America, and eleven US cultivars, included as checks, were evaluated for yield and kernel characteristics, and genotyped with 123 SSR markers. The SSR markers were highly polymorphic across all accessions. Population structure analysis identified eight main clusters for the accessions which corresponded to the major geographic regions, indicating agreement between genetic and predefined populations. Linkage disequilibrium (LD) patterns and distributions are of fundamental importance for genome-wide mapping association. LD between linked markers decreased with distance and with a substantial drop in LD decay values between 20 and 30 cM, suggesting it should be possible to achieve resolution down to the 25 cM level. For the 103 cultivars, the complex traits yield, kernel width, kernel length, kernel width/length ratio, and 1000-kernel weight, were estimated by analysis of variety trial data. The mixed linear model method was used to disclose marker-trait associations. Many of the associated markers were located in regions where QTL had previously been identified. In conclusion, association mapping in rice is a viable alternative to QTL mapping based on crosses between different lines.

/pdf/association-mapping-of-yield-and-its-components-in-rice-b4crrl1nty.pdf

Association mapping of yield and its components in rice cultivars

Association analysis is a method potentially useful for detection of marker-trait associations based on linkage disequilibrium, but little information is available on the application of this technique to plant breeding populations. With appropriate statistical methods, valid association analysis can be done in plant breeding populations; however, the most significant marker may not be closest to the functional gene. Bias can arise from (i) covariance among markers and QTL, frequently related to population structure or intense selection and (ii) differences in initial frequencies of marker alleles in the population, such that exclusive alleles tend to be in higher association. The potentials and limitations of germplasm bank collections, synthetic populations, and elite germplasm are compared, as experimental materials for association analysis integrated with plant breeding practice. Synthetics offer a favorable balance of power and precision for association analysis and would allow mapping of quantitative traits with increasing resolution through cycles of intermating. A model to describe the association between markers and genes as conditional probabilities in synthetic populations under recurrent selection is proposed, which can be computed on the basis of assumptions related to the history of the population. This model is useful for predicting the potential of different populations for association analysis and forecasting the response to marker-assisted selection.

Association Analysis as a Strategy for Improvement of Quantitative Traits in Plants

Simple sequence repeats (SSRs) in DNA have been traditionally thought of as functionally unimportant and have been studied mainly as genetic markers. A recent handful of studies have shown, however, that SSRs in different positions of a gene can play important roles in determining protein function, genetic development, and regulation of gene expression. We have performed a detailed comparative study of the distribution of SSRs in the sequenced genomes of Arabidopsis thaliana and rice. SSRs in different genic regions - 5'untranslated region (UTR), 3'UTR, exon, and intron - show distinct patterns of distribution both within and between the two genomes. Especially notable is the much higher density of SSRs in 5'UTRs compared to the other regions and a strong affinity towards trinucleotide repeats in these regions for both rice and Arabidopsis. On a genomic level, mononucleotide repeats are the most prevalent type of SSRs in Arabidopsis and trinucleotide repeats are the most prevalent type in rice. Both plants have the same most common mononucleotide (A/T) and dinucleotide (AT and AG) repeats, but have little in common for the other types of repeats. Our work provides insight into the evolution and distribution of SSRs in the two sequenced model plant genomes of monocots and dicots. Our analyses reveal that the distributions of SSRs appear highly non-random and vary a great deal in different regions of the genes in the genomes.

/pdf/distinct-patterns-of-ssr-distribution-in-the-arabidopsis-t43k8381f5.pdf

Distinct patterns of SSR distribution in the Arabidopsis thaliana and rice genomes

The archipelago of Indonesia has a long history of rice production across a broad range of rice-growing environments resulting in a diverse array of local Indonesian rice varieties. Although some have been incorporated into modern breeding programs, the vast majority of these landraces remain untapped. To better understand this rich source of genetic diversity we have characterized 330 rice accessions, including 246 Indonesian landraces and 63 Indonesian improved cultivars, using 30 fluorescently-labeled microsatellite markers. The landraces were selected across 21 provinces and include representatives of the classical subpopulations of cere, bulu, and gundil rices. A total of 394 alleles were detected at the 30 simple sequence repeat loci, with an average number of 13 alleles per locus across all accessions, and an average polymorphism information content value of 0.66. Genetic diversity analysis characterized the Indonesian landraces as 68% indica and 32% tropical japonica, with an indica gene diversity of 0.53 and a tropical japonica gene diversity of 0.56, and a Fst of 0.38 between the two groups. All of the improved varieties sampled were indica, and had an average gene diversity of 0.46. A set of high quality Indonesian varieties, including Rojolele, formed a separate cluster within the tropical japonicas. This germplasm presents a valuable source of diversity for future breeding and association mapping efforts.

Genetic diversity analysis of traditional and improved Indonesian rice (Oryza sativa L.) germplasm using microsatellite markers.

This study was undertaken to investigate the population structure of U.S. rice (Oryza sativa L.). A total of 115 U.S. rice cultivars and 30 ancestral accessions introduced from Asia were genotyped by means of 169 simple sequence repeat (SSR) markers that are well distributed throughout the rice genome. SSR-based clustering analysis identified three groups of U.S. rice cultivars that were recognizable as temperate japonica with short to medium grains, tropical japonica with medium grains, and tropical japonica with long grains. Indica cultivars were represented among ancestral accessions, but always clustered independently. Indica germplasm has been used for cultivar improvement, but never directly in U.S. rice production. Cluster analysis of cultivars based on four time periods representing their first release date or introduction (1900-1929, 1930-1959, 1960-1979, and 1980-2000) resulted in the identification of the same three groups. This suggests that the population structure in U.S. rice was established before 1930 and remains essentially intact today, despite a large amount of controlled crossing and artificial selection as a part of the breeding process. Fifty-seven percent of U.S. rice cultivars were developed from intragroup crosses, indicating the availability of substantial genetic variability within each group. Similar results were obtained using genetic distance-based and model-based clustering methods. Information about population structure and associated phenotypic characteristics recognized by geneticists and breeders paves the way for coordinated association mapping studies in the future.

Population Structure and Breeding Patterns of 145 U.S. Rice Cultivars Based on SSR Marker Analysis

The Pi-ta gene from indica introgressed into japonica rice has been used to control the blast disease caused by the fungal pathogen Magnaporthe grisea (Herbert) Barr. (anamorph Pyricularia oryzae Cav.) worldwide. A single nucleotide length polymorphism (SNLP) was identified at the intron region of the Pi-ta gene to develop a codominant Pi-ta gene marker suitable for genotyping with an automated machine. The DNA primer specific to the resistant Pi-taallele was labeled with blue dye (FAM, 6-carboxyfluorescein) as a forward primer, the DNA primer specific to the susceptible pi-ta allele was labeled with green dye (HEX, 4,7,2′,4′,5′,7′-hexachloro-6-carboxyfluorescein) as another forward primer and the DNA primer identical to both Pi-ta/pi-ta alleles was unlabeled as the reverse primer for polymerase chain reaction (PCR). Using these three primers, a 181-bp blue peak in homozygous resistant and a green peak of 182–183 bp in homozygous susceptible, and both peaks in heterozygous plants were produced by PCR. The utility of marker was verified using a segregating F2 population, inbred cultivated lines, dominant markers and pathogenicity testing. A codominant Pi-ta marker was thus developed for effective Pi-ta assisted selection for crop improvement. Using highly homologous competitive primers for allele detection by PCR can benefit the study of genome organization of the complex locus.

Marc A. Redus

Papers

Population Structure and Breeding Patterns of 145 U.S. Rice Cultivars Based on SSR Marker Analysis

Development of a SNLP marker from the Pi-ta blast resistance gene by tri-primer PCR