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We previously identified a cluster of yield-related quantitative trait loci (QTLs) including plant height in CR4379, a near-isogenic line from a cross between Oryza sativa spp. japonica cultivar ‘Hwaseong’ and the wild relative Oryza rufipogon. Map-based cloning and transgenic approaches revealed that APX9, which encodes an l-ascorbate peroxidase 4, is associated with this cluster. A 3 bp InDel was observed leading to the addition of a valine in Hwaseong compared with O. rufipogon. APX9-overexpressing transgenic plants in the Hwaseong background were taller than Hwaseong. Consistent with these results, APX9 T-DNA insertion mutants in the japonica cultivar Dongjin were shorter. These results confirm that APX9 is the causal gene for the QTL cluster. Sequence analysis of APX9 from 303 rice accessions revealed that the 3 bp InDel clearly differentiates japonica (APX9HS) and O. rufipogon (APX9OR) alleles. indica accessions shared both alleles, suggesting that APX9HS was introgressed into indica followed by crossing. The finding that O. rufipogon accessions with different origins carry APX9OR suggests that the 3 bp insertion was specifically selected in japonica during its domestication. Our findings demonstrate that APX9 acts as a major regulator of plant development by controlling a valuable suite of agronomically important traits in rice., DNA marker-based smart breeding revealed that the ascorbate peroxidase gene APX9 of the wild rice relative Oryza rufipogon with low yield can make cultivated rice more productive and stress-tolerant.

Genetic analysis of genes that regulate the color pigmentation of sterile lemma and apiculus has been conducted. “Josaengjado” has small and round grains with purple leaf, sterile lemma and apiculus. In the F2 population from a cross between Josaengjado and Daeribbyeo 1, 246 and 182 plants exhibited purple and straw-white sterile lemma, respectively. It fitted a 9:7 segregation ratio indicating that two complementary genes control the pigmentation in sterile lemma and apiculus. Genetic analysis was performed using the F2:3 and KASP (Kompetitive Allele-Specific PCR) markers. Genes for the coloration of leaf sheath, ligule, sterile lemma, and apiculus were detected on chromosomes 1 and 6. Sequence comparison showed a single nucleotide substitution C (Josaengjado) to A (Daeribbyeo 1) in the second exon of the Rd gene on chromosome 1 leading to a premature stop in Daeribbyeo 1. In C1, a 3-bp deletion in the second exon was detected in Daeribbyeo 1. Haplotype analysis was performed in the Rd and C1 genes of the 78 rice accessions. 78 accessions were divided into 14 groups. A total of 11 and 1 mutation sites were detected in OsC1 and Rd, respectively. The haplotype analysis also confirmed that two complementary genes, Rd and OsC1 are necessary to express anthocyanin pigmentation in sterile lemma and apiculus. To our knowledge, this is the first report to identify genes for the coloration of sterile lemma in rice.

Low-temperature germinability (LTG) is an important trait for rice direct seeding at temperate and high-altitude region. Previously, five QTLs (quantitative trait loci) for LTG were detected using an interspecific cross population between the Korea japonica cultivar Hwaseong and Oryza rufipogon (IRGSP#105491). O. rufipogon alleles at all loci increased the germination rate at the 13℃ condition. In this study, we tried to confirm and fine-map qLTG1 located on the short arm of chromosome 1. To map the qLTG1, two introgression lines, TR5 and TR20 were crossed to Hwaseong to develop F2:3 populations. QTL analysis confirmed the existence of the qLTG1 and it explained 55.5% and 29.9% of phenotypic variation in two populations, respectively. Substitution mapping using informative recombinant lines indicated that qLTG1 was located in 167-kb region between two SSR markers RM10310 and RM10326. This segment harbored 18 genes with nine of them were annotated with specific gene function. In addition, the O. rufipogon introgression in this region was associated with an increase in spikelets per panicle in the Hwaseong background. The results strongly indicate that the O. rufipogon alleles will be a valuable source of genes in improving japonica rice for low-temperature germinability and yield. To our knowledge, this is the first report to fine-map qLTG1 associated with LTG in rice considering that no QTL for LTG has not been reported near this QTL region from other biparental populations.

BackgroundRice (Oryza sativaL.) is generally sensitive to low temperatures, and in production systems that use direct-seeding, low-temperature germinability (LTG) is a desired trait. Previously, the QTLs,qLTG1andqLTG3,that control LTG, were mapped using the BC4F8population, which is a cross of Korean elite cultivar Hwaseong andO. rufipogon(IRGC 105491). We have characterized and analyzed the interaction between the two QTLs, by crossing TR20 that hasO. rufipogonalleles atqLTG1andqLTG3in a Hwaseong background, with Hwaseong, to develop an F2population.ResultsThe F2plants with bothqLTG1andqLTG3alleles fromO. rufipogonshowed higher LTG scores, than the plants with onlyqLTG1orqLTG3. No significant interaction between theqLTG1andqLTG3was observed, indicating that they may regulate LTG via different pathways. Based on its location,qLTG3appears to be allelic withqLTG3–1, a major QTL known to control LTG. To investigate the genetic differences between the two parents, that were controlling LTG, we compared theirqLTG3–1sequences. In the coding region, three sequence variations leading to amino acid changes were identified between the Hwaseong andO. rufipogon. Of these, a non-synonymous substitution at the 62nd amino acid site, had not previously been reported. To understand the cause of the LTG variations between the parents, we genotyped three sequence variations ofqLTG3–1, that were identified in 98 Asian cultivated rice accessions (Oryza sativaL.). The 98 accessions were classified into 5 haplotypes, based on three variations and a 71-bp deletion. Mean low-temperature germination rates were compared among the haplotypes, and haplotype 5 (O. rufipogon-type) showed a significantly higher germination rate than haplotype 2 (Nipponbare-type), and haplotype 3 (Italica Livorno-type).ConclusionsTheO. rufipogon qLTG3–1allele can be utilized for the improvement of LTG in rice breeding programs. Nearly isogenic lines harboring bothqLTG1andqLTG3–1alleles fromO. rufipogon,showed higher LTG scores than the NILs withqLTG1orqLTG3–1alone, and the two QTLs regulate LTG via different pathways. To our knowledge, this is the first report to detect a newqLTG3–1allele and analyze the interaction of the two LTG QTLs in a nearly isogenic background.

A quantitative trait locus (QTL) gw8.1 was detected in the population derived from a cross between the elite japonica cultivar, 'Hwaseong' and Oryza rufipogon (IRGC 105491). Near isogenic lines (NILs) harboring the O. rufipogon segment on chromosome 8 showed increased grain length and weight compared to those of the recurrent parent, Hwaseong. This QTL was mapped to a 175.3-kb region containing 28 genes, of which four were considered as candidates based on the presence of mutations in their coding regions and as per the RNA expression pattern during the inflorescence stage. Leaves and panicles obtained from plants harvested 5 days after heading showed differences in gene expression between Hwaseong and gw8.1-NILs. Most genes were upregulated in O. rufipogon and gw8.1-NIL than in Hwaseong. Scanning electron microscopy analysis of the lemma inner epidermal cells indicated that cell length was higher in gw8.1 NIL than in Hwaseong, indicating that gw8.1 might regulate cell elongation. Among the candidate genes, LOC_Os08g34380 encoding a putative receptor-like kinase and LOC_Os08g34550 encoding putative RING-H2 finger protein were considered as possible candidates based on their functional similarity.

Previously, five putative quantitative trait loci (QTLs) for low-temperature germination (LTG) have been detected using 96 BC3F8 lines derived from an interspecific cross between the Korean japonica cultivar &ldquo, Hwaseong&rdquo, and Oryza rufipogon. In the present study, two introgression lines, CR1517 and CR1518, were used as parents to detect additional QTLs and analyze interactions among QTLs for LTG. The F2 population (154 plants) along with parental lines, Hwaseong and O. rufipogon, were evaluated for LTG and coleoptile length under low-temperature conditions (13 &deg, C). Among five QTLs for LTG, two major QTLs, qLTG1 and qLTG3, were consistently detected at 6 and 7 days after incubation. Three minor QTLs were detected on chromosomes 8 and 10. Two QTLs, qLTG10.1 and qLTG10.2, showing linkage on chromosome 10, exerted opposite effects with the Hwaseong allele at qLTG10.2 and the O. rufipogon allele at qLTG10.1 respectively, in turn, increasing LTG. Interactions among QTLs were not significant, implying that the QTLs act in an additive manner. Near-isogenic line plants with the combination of favorable alleles from O. rufipogon and Hwaseong exhibited higher LTG than two introgression lines. With regard to coleoptile length, three QTLs observed on chromosomes 1, 3, and 8 were colocalized with QTLs for LTG, suggesting the pleiotropy of the single gene at each locus. According to the results, the introgression of favorable O. rufipogon alleles could hasten the development of rice with high LTG and high coleoptile elongation in japonica cultivars.