Amy Bernardo

Discovery and mapping of single feature polymorphisms in wheat using Affymetrix arrays

BackgroundWheat (Triticum aestivum L.) is a staple food crop worldwide. The wheat genome has not yet been sequenced due to its huge genome size (~17,000 Mb) and high levels of repetitive sequences; the whole genome sequence may not be expected in the near future. Available linkage maps have low marker density due to limitation in available markers; therefore new technologies that detect genome-wide polymorphisms are still needed to discover a large number of new markers for construction of high-resolution maps. A high-resolution map is a critical tool for gene isolation, molecular breeding and genomic research. Single feature polymorphism (SFP) is a new microarray-based type of marker that is detected by hybridization of DNA or cRNA to oligonucleotide probes. This study was conducted to explore the feasibility of using the Affymetrix GeneChip to discover and map SFPs in the large hexaploid wheat genome.ResultsSix wheat varieties of diverse origins (Ning 7840, Clark, Jagger, Encruzilhada, Chinese Spring, and Opata 85) were analyzed for significant probe by variety interactions and 396 probe sets with SFPs were identified. A subset of 164 unigenes was sequenced and 54% showed polymorphism within probes. Microarray analysis of 71 recombinant inbred lines from the cross Ning 7840/Clark identified 955 SFPs and 877 of them were mapped together with 269 simple sequence repeat markers. The SFPs were randomly distributed within a chromosome but were unevenly distributed among different genomes. The B genome had the most SFPs, and the D genome had the least. Map positions of a selected set of SFPs were validated by mapping single nucleotide polymorphism using SNaPshot and comparing with expressed sequence tags mapping data.ConclusionThe Affymetrix array is a cost-effective platform for SFP discovery and SFP mapping in wheat. The new high-density map constructed in this study will be a useful tool for genetic and genomic research in wheat.

Single nucleotide polymorphism in wheat chromosome region harboring Fhb1 for Fusarium head blight resistance

Fusarium head blight (FHB) is a destructive disease that reduces wheat grain yield and quality. To date, the quantitative trait locus on 3BS (Fhb1) from Sumai 3 has shown the largest effect on FHB resistance. Single nucleotide polymorphism (SNP) is the most common form of genetic variation and is suitable for high-throughput marker-assisted selection (MAS). We analyzed SNPs derived from 23 wheat expressed sequence tags (ESTs) that previously mapped near Fhb1 on chromosome 3BS. Using 71 Ning 7840/Clark BC7F7 recombinant inbred lines and the single-base extension method, we mapped seven SNP markers between Xgwm533 and Xgwm493, flanking markers for Fhb1. Five of the SNPs explained 45–54% of the phenotypic variation for FHB resistance. Haplotype analysis of 63 wheat accessions from eight countries based on SNPs in EST sequences, simple sequence repeats, and sequence tagged sites in the Fhb1 region identified four major groups: (1) US-Clark, (2) Asian, (3) US-Ernie, and (4) Chinese Spring. The Asian group consisted of Chinese and Japanese accessions that carry Fhb1 and could be differentiated from other groups by marker Xsnp3BS-11. All Sumai 3-related accessions formed a subgroup within the Asian group and could be sorted out by Xsnp3BS-8. The SNP markers identified in this study should be useful for MAS of Fhb1 and fine mapping to facilitate cloning of the Fhb1 resistance gene.

Adult plant leaf rust resistance derived from toropi wheat is conditioned by Lr78 and three minor QTL

Leaf rust caused by Puccinia triticina is an important disease of wheat in many regions worldwide. Durable or long-lasting leaf rust resistance has been difficult to achieve because populations of P. triticina are highly variable for virulence to race-specific resistance genes, and respond to selection by resistance genes in released wheat cultivars. The wheat cultivar Toropi, developed and grown in Brazil, was noted to have long-lasting leaf rust resistance that was effective only in adult plants. The objectives of this study were to determine the chromosome location of the leaf rust resistance genes derived from Toropi in two populations of recombinant inbred lines in a partial Thatcher wheat background. In the first population, a single gene with major effects on chromosome 5DS that mapped 2.2 centimorgans distal to IWA6289, strongly reduced leaf rust severity in all 3 years of field plot tests. This gene for adult plant leaf rust resistance was designated as Lr78. In the second population, quantitative trait loci (QTL) with small effects on chromosomes 1BL, 3BS, and 4BS were found. These QTL expressed inconsistently over 4 years of field plot tests. The adult plant leaf rust resistance derived from Toropi involved a complex combination of QTL with large and small effects.

Mapping and characterization of the new adult plant leaf rust resistance gene Lr77 derived from Santa Fe winter wheat

Key messageA new gene for adult plant leaf rust resistance in wheat was mapped to chromosome 3BL. This gene was designated as Lr77.Abstract‘Santa Fe’ is a hard red winter cultivar that has had long-lasting resistance to the leaf rust fungus, Puccinia triticina. The objective of this study was to determine the chromosome location of the adult plant leaf rust resistance in Santa Fe wheat. A partial backcross line of ‘Thatcher’ (Tc) wheat with adult plant leaf rust resistance derived from Santa Fe was crossed with Thatcher to develop a Thatcher//Tc*2/Santa Fe F6 recombinant inbred line (RIL) population. The RIL population and parental lines were evaluated for segregation of leaf rust resistance in three field plot tests and in an adult plant greenhouse test. A genetic map of the RIL population was constructed using 90,000 single-nucleotide polymorphism (SNP) markers with the Illumina Infinium iSelect 90K wheat bead array. A significant quantitative trait locus for reduction of leaf rust severity in all four tests was found on chromosome 3BL that segregated as a single adult plant resistance gene. The RILs with the allele from the resistant parent for SNP marker IWB10344 had lower leaf rust severity and a moderately resistant to moderately susceptible response compared to the susceptible RILs and Thatcher. The gene derived from Santa Fe on chromosome 3BL was designated as Lr77. Kompetitive allele-specific polymerase chain reaction assay markers linked to Lr77 on 3BL should be useful for selection of wheat germplasm with this gene.