As an essential macroelement for all living cells, phosphorus is indispensable in agricultural production systems. Natural phosphorus reserves are limited, and it is therefore important to develop phosphorus-efficient crops. A major quantitative trait locus for phosphorus-deficiency tolerance, Pup1, was identified in the traditional aus-type rice variety Kasalath about a decade ago. However, its functional mechanism remained elusive until the locus was sequenced, showing the presence of a Pup1-specific protein kinase gene, which we have named phosphorus-starvation tolerance 1 (PSTOL1). This gene is absent from the rice reference genome and other phosphorus-starvation-intolerant modern varieties. Here we show that overexpression of PSTOL1 in such varieties significantly enhances grain yield in phosphorus-deficient soil. Further analyses show that PSTOL1 acts as an enhancer of early root growth, thereby enabling plants to acquire more phosphorus and other nutrients. The absence of PSTOL1 and other genes-for example, the submergence-tolerance gene SUB1A-from modern rice varieties underlines the importance of conserving and exploring traditional germplasm. Introgression of this quantitative trait locus into locally adapted rice varieties in Asia and Africa is expected to considerably enhance productivity under low phosphorus conditions.

The major quantitative trait locus (QTL) Phosphorus uptake1 (Pup1) confers tolerance of phosphorus deficiency in soil and is currently one of the most promising QTLs for the development of tolerant rice (Oryza sativa) varieties. To facilitate targeted introgression of Pup1 into intolerant varieties, the gene models predicted in the Pup1 region in the donor variety Kasalath were used to develop gene-based molecular markers that are evenly distributed over the fine-mapped 278-kb QTL region. To validate the gene models and optimize the markers, gene expression analyses and partial allelic sequencing were conducted. The markers were tested in more than 80 diverse rice accessions revealing three main groups with different Pup1 allele constitution. Accessions with tolerant (group I) and intolerant (group III) Pup1 alleles were distinguished from genotypes with Kasalath alleles at some of the analyzed loci (partial Pup1; group II). A germplasm survey additionally confirmed earlier data showing that Pup1 is largely absent from irrigated rice varieties but conserved in varieties and breeding lines adapted to drought-prone environments. A core set of Pup1 markers has been defined, and sequence polymorphisms suitable for singlenucleotide polymorphism marker development for high-throughput genotyping were identified. Following a marker-assisted backcrossing approach, Pup1 was introgressed into two irrigated rice varieties and three Indonesian upland varieties. First phenotypic evaluations of the introgression lines suggest that Pup1 is effective in different genetic backgrounds and environments and that it has the potential to significantly enhance grain yield under field conditions.

Marker-assisted breeding is a very useful tool for breeders but still lags behind its potential because information on the effect of quantitative trait loci (QTLs) in different genetic backgrounds and ideal molecular markers are unavailable. Here, we report on some first steps toward the validation and application of the major rice QTL Phosphate uptake 1 (Pup1) that confers tolerance of phosphorus (P) deficiency in rice (Oryza sativa L.). Based on the Pup1 genomic sequence of the tolerant donor variety Kasalath that recently became available, markers were designed that target (1) putative genes that are partially conserved in the Nipponbare reference genome and (2) Kasalath-specific genes that are located in a large insertion-deletion (INDEL) region that is absent in Nipponbare. Testing these markers in 159 diverse rice accessions confirmed their diagnostic value across genotypes and showed that Pup1 is present in more than 50% of rice accessions adapted to stress-prone environments, whereas it was detected in only about 10% of the analyzed irrigated/lowland varieties. Furthermore, the Pup1 locus was detected in more than 80% of the analyzed drought-tolerant rice breeding lines, suggesting that breeders are unknowingly selecting for Pup1. A hydroponics experiment revealed genotypic differences in the response to P deficiency between upland and irrigated varieties but confirmed that root elongation is independent of Pup1. Contrasting Pup1 near-isogenic lines (NILs) were subsequently grown in two different P-deficient soils and environments. Under the applied aerobic growth conditions, NILs with the Pup1 locus maintained significantly higher grain weight plant(-1) under P deprivation in comparison with intolerant sister lines without Pup1. Overall, the data provide evidence that Pup1 has the potential to improve yield in P-deficient and/or drought-prone environments and in diverse genetic backgrounds.

To identify quantitative trait loci (QTL) controlling heat tolerance in rice, the progeny of BC 1 F 1 and F 2 populations derived from an IR64 · N22 cross were exposed to 38/24°C for 14 days at the flowering stage, and spikelet fertility was assessed. A custom 384-plex Illumina GoldenGate genotyping assay was used to genotype the F 2 and selected BC 1 F 1 plants. Four single nucleotide polymorphisms were associated with heat tolerance in the BC 1 F 1 population using selective genotyping and single marker analysis, and four putative QTL were found to be associated with heat tolerance in the F 2 population. Two major QTL were located on chromosome 1 (qHTSF1.1) and chromosome 4 (qHTSF4.1). These two major QTL could explain 12.6% (qHTSF1.1) and 17.6% (qHTSF4.1) of the variation in spikelet fertility under high temperature. Tolerant allele of qHTSF1.1 was from the susceptible parent IR64, and that of qHTSF4.1 was from tolerant parent N22. The effect of qHTSF4.1 on chromosome 4 was confirmed in selected BC 2 F 2 progeny from the same IR64 · N22 cross, and the plants with qHTSF4.1 showed significantly higher spikelet fertility than other genotypes.

Brown planthopper (BPH) is one of the most destructive insect pests of rice. Wild species of rice are a valuable source of resistance genes for developing resistant cultivars. A molecular marker-based genetic analysis of BPH resistance was conducted using an F(2) population derived from a cross between an introgression line, 'IR71033-121-15', from Oryza minuta (Accession number 101141) and a susceptible Korean japonica variety, 'Junambyeo'. Resistance to BPH (biotype 1) was evaluated using 190 F(3) families. Two major quantitative trait loci (QTLs) and two significant digenic epistatic interactions between marker intervals were identified for BPH resistance. One QTL was mapped to 193.4-kb region located on the short arm of chromosome 4, and the other QTL was mapped to a 194.0-kb region on the long arm of chromosome 12. The two QTLs additively increased the resistance to BPH. Markers co-segregating with the two resistance QTLs were developed at each locus. Comparing the physical map positions of the two QTLs with previously reported BPH resistance genes, we conclude that these major QTLs are new BPH resistance loci and have designated them as Bph20(t) on chromosome 4 and Bph21(t) on chromosome 12. This is the first report of BPH resistance genes from the wild species O. minuta. These two new genes and markers reported here will be useful to rice breeding programs interested in new sources of BPH resistance.

SummaryA rice genic male-sterility gene ms-h is recessive and has a pleiotropic effect on the chalky endosperm. After fine mapping, nucleotide sequencing analysis of the ms-h gene revealed a single nucleotide substitution at the 3¢-splice junction of the 14th intron of the UDP-glucose pyrophosphorylase 1 (UGPase1; EC2.7.7.9) gene, which causes the expression of two mature transcripts with abnormal sizes caused by the aberrant splicing. An in vitro functional assay showed that both proteins encoded by the two abnormal transcripts have no UGPase activity. The suppression of UGPase by the introduction of a UGPase1-RNAi construct in wild-type plants nearly eliminated seed set because of the male defect, with developmental retardation similar to the ms-h mutant phenotype, whereas overexpression of UGPase1 in ms-h mutant plants restored male fertility and the transformants produced T 1 seeds that segregated into normal and chalky endosperms. In addition, both phenotypes were co-segregated with the UGPase1 transgene in segregating T 1 plants, which demonstrates that UGPase1 has functional roles in both male sterility and the development of a chalky endosperm. Our results suggest that UGPase1 plays a key role in pollen development as well as seed carbohydrate metabolism.

SummaryThe p hosphorus up take 1 ( Pup1 ) locus was identified as a major quantitative trait locus (QTL) for tolerance of phosphorus deficiency in rice. Near-isogenic lines with the Pup1 region from tolerant donor parent Kasalath typically show threefold higher phosphorus uptake and grain yield in phosphorus-deficient field trials than the intolerant parent Nipponbare. In this study, we report the fine mapping of the Pup1 locus to the long arm of chromosome . Genes in the region were initially identified on the basis of the Nipponbare reference genome, but did not reveal any obvious candidate genes related to phosphorus uptake. Kasalath BAC clones were therefore sequenced and revealed a 278-

BackgroundFixed arrays of single nucleotide polymorphism (SNP) markers have advantages over reduced representation sequencing in their ease of data analysis, consistently higher call rates, and rapid turnaround times. A 6 K SNP array represents a cost-benefit “sweet spot” for routine genetics and breeding applications in rice. Selection of informative SNPs across species and subpopulations during chip design is essential to obtain useful polymorphism rates for target germplasm groups. This paper summarizes results from large-scale deployment of an Illumina 6 K SNP array for rice.ResultsDesign of the Illumina Infinium 6 K SNP chip for rice, referred to as the Cornell_6K_Array_Infinium_Rice (C6AIR), includes 4429 SNPs from re-sequencing data and 1571 SNP markers from previous BeadXpress 384-SNP sets, selected based on polymorphism rate and allele frequency within and between target germplasm groups. Of the 6000 attempted bead types, 5274 passed Illumina’s production quality control. The C6AIR was widely deployed at the International Rice Research Institute (IRRI) for genetic diversity analysis, QTL mapping, and tracking introgressions and was intensively used at Cornell University for QTL analysis and developing libraries of interspecific chromosome segment substitution lines (CSSLs) between O. sativa and diverse accessions of O. rufipogon or O. meridionalis. Collectively, the array was used to genotype over 40,000 rice samples. A set of 4606 SNP markers was used to provide high quality data for O. sativa germplasm, while a slightly expanded set of 4940 SNPs was used for O. sativa X O. rufipogon populations. Biparental polymorphism rates were generally between 1900 and 2500 well-distributed SNP markers for indica x japonica or interspecific populations and between 1300 and 1500 markers for crosses within indica, while polymorphism rates were lower for pairwise crosses within U.S. tropical japonica germplasm. Recently, a second-generation array containing ~7000 SNP markers, referred to as the C7AIR, was designed by removing poor-performing SNPs from the C6AIR and adding markers selected to increase the utility of the array for elite tropical japonica material.ConclusionsThe C6AIR has been successfully used to generate rapid and high-quality genotype data for diverse genetics and breeding applications in rice, and provides the basis for an optimized design in the C7AIR.Electronic supplementary materialThe online version of this article (10.1186/s12284-017-0181-2) contains supplementary material, which is available to authorized users.