IC4R008-GWAS-2016-27228161

Project Title

Genome-Wide Association Study for Traits Related to Plant and Grain Morphology, and Root Architecture in Temperate Rice Accessions

In this study the researchers carried out a genome-wide association analysis for plant and grain morphology and root architecture in a unique panel of temperate rice accessions adapted to European pedo-climatic conditions. This is the first study to assess the association of selected phenotypic traits to specific genomic regions in the narrow genetic pool of temperate japonica. A set of 391 rice accessions were GBS-genotyped yielding—after data editing—57000 polymorphic and informative SNPS, among which 54% were in genic regions.

The accession panel used in this study included 391 Oryza sativa varieties from the Rice Germplasm Collection maintained at the CREA-Rice Research Unit (Vercelli, Italy). Varieties were selected based on the results of a genetic diversity analysis with 24 independent SSR markers; the aim was to include the broadest possible range of genetically and phenotypically diverse temeperate rice accessions.

The sampled collection included 9 aromatic, 11 aus, 22 indica, 96 tropical japonica and 253 temperate japonica accessions. Most of these were temperate rice developed and selected in Italy (126 accessions), or developed elsewhere and adapted to Italian agro-climatic conditions (127 accessions). This collection was representative of the worldwide rice diversity, and was used to evaluate the population structure of the analysed rice accession panel.

The distribution of the 391 accessions per taxonomic group can be seen in Table 1. The complete list of the accessions used in this study, with information on taxonomic group and geographic origin is reported in S1 Table. All accessions were purified through single seed descent before genotyping and phenotypic evaluation. From all 391 accessions DNA was extracted for genotyping; phenotypes were collected on a subset of temperate and tropical japonica accessions: 153 for plant morphology and grain related traits, and 93 for root traits (Table 1).

Table 1 Taxonomic classification of accessions.

Total genomic DNA was isolated from three-week old leaves using the DNeasy Plant Mini Kit (QIAGEN) with a TECAN Freedom EVO150 liquid handling robot (TECAN Group Ltd, Switzerland). For each accession, a single individual plant was used.The libraries were loaded into Genome Analyzer II (Illumina, Inc., San Diego, CA) for sequencing. Raw sequence data filtering, sequence alignment to the rice reference genome (Os-Nipponbare-Reference-IRGSP-1.0,) and SNP calling from low-coverage GBS genotyping, were carried out using the Tassel GBS pipeline v3.0 provided by the Buckler Lab for Maize Genetics and Diversity。 A total of 166418 SNPs were called from GBS genotyping. This information is eventually used to impute the target locus in an iterative approach that attemps, through a Markov Chain MonteCarlo (MCMC) process, to maximise the likelihood of the observed SNP calls given the unobserved imputed genotypes.

In total, 42 significant genotype-phenotype associations were detected: 21 for plant morphology traits, 11 for grain quality traits, 10 for root architecture traits.

The FDR of detected associations ranged from 3·10−7 to 0.92 (median: 0.25). In most cases, the significant detected associations co-localised with QTLs and candidate genes controlling the phenotypic variation of single or multiple traits. The most significant associations were those for flag leaf width on chromosome 4 (FDR = 3·10−7) and for plant height on chromosome 6 (FDR = 0.011).

The researchers demonstrate the effectiveness and resolution of the developed platform for highthroughput phenotyping, genotyping and GWAS in detecting major QTLs for relevant traits in rice. The researchers identified strong associations that may be used for selection in temperate irrigated rice breeding: e.g. associations for flag leaf width, plant height, root volume and length, grain length, grain width and their ratio.

PTP Science Park, 26900 Lodi, Italy
IBBA-CNR, Lodi, Italy
CREA-Council for Agricultural Research and Economics, Rice Research Unit, 13100 Vercelli, Italy
CREA-Council for Agricultural Research and Economics, Genomics Research Centre, 29017 Fiorenzuola d’Arda (Piacenza), Italy, 5 Department of Plant Breeding & Genetics, Cornell University, Ithaca, NY, United States of America