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The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a major insect pest of cultivated rice, Oryza sativa L., throughout the temperate regions of East Asia. GRH resistance had been reported in the wild species Oryza nivara but genetic basis of GRH resistance in wild rice accession has not been clarified. Here, we found a major QTL, qGRH4.2, on chromosome 4 conferred GRH resistance with 14.1 of the logarithm of odds (LOD) score explaining 67.6% of phenotypic variance in the BC1F1 population derived from a cross between the susceptible japonica cultivar ‘Taichung 65’ (T65) and O. nivara accession IRGC105715. qGRH4.2 has been identified as GRH6 between the markers RM5414 and C60248 in a BC3F2 population derived from two BC3F1 plants resistant to GRH. In a high-resolution mapping, the GRH6 region was delimited between the markers G6-c60k and 7L16f, and corresponded to an 31.2-kbp region of the ‘Nipponbare’ genome. Understanding the genetic basis of GRH resistance will facilitate the use of GRH resistance genes in marker-assisted breeding in rice.

To evaluate and utilize potentially valuable quantitative trait loci or genes of wild relatives in the genetic background of domesticated crop species, chromosome segment substitution lines (CSSLs) are a valuable tool. CSSLs can be constructed through the exchange of chromosome segments of AA genome species of the genus Oryza with cultivated rice, Oryza sativa L. Here we report the development of three sets of CSSLs carrying segments of AA genome species closely related to Oryza sativa-O. glaberrima (IRGC 103777 from Mali), O. rufipogon (W1962 from China), and O. nivara (IRGC 105715 from Cambodia)-in the genetic background of ssp. japonica cultivar Taichung 65 through the use of 101 to 121 simple-sequence-repeat markers in whole-genome genotyping and marker-assisted selection. The materials are available via the National Bioresource Project (Rice) Oryzabase Web page.

The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a major insect pest of cultivated rice, Oryza sativa L., throughout the temperate regions of East Asia. GRH resistance had been reported in the wild species Oryza nivara but genetic basis of GRH resistance in wild rice accession has not been clarified. Here, we found a major QTL, qGRH4.2, on chromosome 4 conferred GRH resistance with 14.1 of the logarithm of odds (LOD) score explaining 67.6% of phenotypic variance in the BC1F1 population derived from a cross between the susceptible japonica cultivar ‘Taichung 65’ (T65) and O. nivara accession IRGC105715. qGRH4.2 has been identified as GRH6 between the markers RM5414 and {"type":"entrez-nucleotide","attrs":{"text":"C60248","term_id":"2418953","term_text":"C60248"}}C60248 in a BC3F2 population derived from two BC3F1 plants resistant to GRH. In a high-resolution mapping, the GRH6 region was delimited between the markers G6-c60k and 7L16f, and corresponded to an 31.2-kbp region of the ‘Nipponbare’ genome. Understanding the genetic basis of GRH resistance will facilitate the use of GRH resistance genes in marker-assisted breeding in rice.

DNA markers are useful for identifying genes and developing new genetic materials for breeding and genetic research. High-resolution melting (HRM) analysis can detect a single nucleotide polymorphism (SNP) in two polymerase chain reaction (PCR) fragments as a melting temperature (Tm) difference without additional experimental steps, such as gel electrophoresis. To design a method for developing reliable HRM markers that discriminate between homozygous alleles containing SNPs, we tested new evaluation indexes related to the thermodynamics of double-stranded DNA to find one that maximizes the difference in Tm values between PCR fragments. We found that differences in the change in Gibbs free energy (ΔG°) correlated with actual differences in Tm values. Optimization of the nearest neighboring nucleotide (NNN) of a SNP by nucleotide substitution in the primer and reducing the size of the PCR fragment both enlarged the actual differences in Tm. The genetic DNA markers we developed by NNN substitution, termed NNNs-HRM markers, could be precisely mapped within soybean chromosomes by linkage analysis. We developed a Perl script pipeline to enable the automatic design of a massive number of NNNs-HRM markers; these scripts are freely available and would be useful for practical breeding programs for other plant species.

The rice cultivar ASD7 (Oryza sativa L. ssp. indica) is resistant to the brown planthopper (BPH; Nilaparvata lugens Stal) and the green leafhopper (Nephotettix virescens Distant). Here, we analyzed multiple genetic resistance to BPH and the green rice leafhopper (GRH; Nephotettix cincticeps Uhler). Using two independent F2 populations derived from a cross between ASD7 and Taichung 65 (Oryza sativa ssp. japonica), we detected two QTLs (qBPH6 and qBPH12) for resistance to BPH and one QTL (qGRH5) for resistance to GRH. Linkage analysis in BC2F3 populations revealed that qBPH12 controlled resistance to BPH and co-segregated with SSR markers RM28466 and RM7376 in plants homozygous for the ASD7 allele at qBPH6. Plants homozygous for the ASD7 alleles at both QTLs showed a much faster antibiosis response to BPH than plants homozygous at only one of these QTLs. It revealed that epistatic interaction between qBPH6 and qBPH12 is the basis of resistance to BPH in ASD7. In addition, qGRH5 controlled resistance to GRH and co-segregated with SSR markers RM6082 and RM3381. qGRH5 is identical to GRH1. Thus, we clarified the genetic basis of multiple resistance of ASD7 to BPH and GRH.

Hybrid incompatibility plays an important role in establishment of post-zygotic reproductive isolation. To unveil genetic basis of hybrid incompatibilities between diverged species of genus Oryza AA genome species, we conducted genetic dissection of hybrid sterility loci, S22(t), which had been identified in backcross progeny derived from Oryza sativa ssp. japonica (recurrent parent) and South American wild rice O. glumaepatula near the end of the short arm of chromosome 2. The S22(t) region was found to be composed of two loci, designated S22A and S22B, that independently induce F1 pollen sterility. Pollen grains containing either of the sterile alleles (S22A-glum (s) or S22B-glum (s) ) were sterile if produced on a heterozygous plant. No transmission of the S22A-glum (s) allele via pollen was observed, whereas a low frequency of transmission of S22B-glum (s) was observed. Cytological analysis showed that the sterile pollen grains caused by S22A could reach the bicellular or tricellular stage, and the nearly-sterile pollen grains caused by S22B could reach the tricellular stage. Our genetic analysis showed repulsion linkage effect is possible to induce strong reproductive barrier by high pollen sterility based on recombination value and transmission ratio of hybrid sterility gene to the progeny was influenced by frequency of competitors on fertilization.

Deepwater rice possesses internode elongation ability to avoid drowning under deepwater conditions. Previous studies identified three QTLs regulating internode elongation ability on chromosomes 1, 3 and 12 using different populations. However, these QTLs only induce internode elongation in response to deepwater conditions from the 7-leaf stage and not during the early leaf stage. In this study, we detected two novel QTLs, qTIL2 and qTIL4 regulating deepwater response at the early leaf stage using an F(2) population derived from the cross between NIL1-3-12 carrying the three QTLs regulating deepwater response in T65 (O. sativa ssp. japonica) genetic background and C9285 (O. sativa ssp. indica, deepwater rice). Plants of the BC(2)F(2) population derived from NIL1-3-12/C9285 and the RILs of T65/Bhadua (O. sativa ssp. indica, deepwater rice) possessing these QTLs as well as the three QTLs previously identified also showed internode elongation during the early leaf stage. These results indicate that qTIL2 and qTIL4 regulate early internode elongation and function in coordination with the three major QTLs under deepwater conditions. The results presented here would not only help define the mechanism of deepwater response in rice but also contribute in the breeding of deepwater tolerant rice that is adapted to various water depths.

Stigma exsertion is an important trait that contributes to the improvement of seed production in hybrid rice. We demonstrate that GS3, one of the genes regulating seed length, also regulates stigma length and participates in stigma exsertion in rice. GS3 mRNA is expressed in the basal part of the young stigma, and a nonsense mutation in the second exon of GS3 causes an increase in cell number, resulting in elongation of the stigma. Manipulation of GS3 should contribute to the improvement of hybrid seed-production efficiency.

The National BioResource Project has been organized and established to promote research activities using valuable bioresources. A total of twenty-eight bioresources for ten animals, nine plants and nine microorganisms/cell lines developed or collected in Japan were selected for the project. Resources are categorized into several different groups in the project; genetic resources, germplasm, genome resources and their information. Choices of how many resources must be preserved and maintained and in which categories are dependent on the status of the research community of each organism. These resources, if utilized systematically and intelligently, are powerful means for leading new scientific discoveries. Some examples can be seen in this paper. This paper reviews plant bioresources with the main focus on rice resource activities within the project.

The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a serious insect pest of cultivated rice (Oryza sativa L.) in temperate East Asia. An African rice cultivar, Oryza glaberrima Steud. (IRGC104038), was shown to be highly GRH-resistant at the booting stage. To reveal the genetic basis of the GRH resistance in O. glaberrima, a BC1F1 population derived from a cross between a susceptible japonica variety, Taichung 65 (T65), and O. glaberrima was analyzed by quantitative trait locus (QTL) analysis. A single major QTL for GRH resistance, designated qGRH9, was detected on rice chromosome 9, and three minor QTLs were detected on rice chromosomes 3, 7, and 10. A series of O. glaberrima introgression lines (GILs) containing IRGC104038 chromosome segments in the T65 genetic background were evaluated to confirm the genetic effects of the QTLs, and three GILs carrying qGRH9 showed resistance to GRH. Substitution mapping using the GILs revealed that qGRH9 was located between simple sequence repeat (SSR) markers RM215 and RM2482 in a 1.39-Mbp region on the distal region of the long arm of chromosome 9, and tightly linked to RM7306. These SSR markers maybe useful for marker-assisted selection of qGRH9 for improvement of GRH resistance in rice.

The brown planthopper [BPH; Nilaparvata lugens (Stal.)] is one of the most destructive insect pests in Asian rice-growing areas. Two genes conferring resistance to BPH, BPH25(t) and BPH26(t) derived from a BPH-resistant indica rice cultivar, Oryza sativa ADR52, have been identified. However, they are linked to genes conferring late heading and hybrid spikelet sterility. To eliminate these unfavorable traits (linkage drag), we generated BC6F1 populations carrying BPH25(t) or BPH26(t) in a BPH-susceptible japonica cultivar, Taichung 65, through marker-assisted selection. We selected three near-isogenic lines (NILs) carrying BPH25(t) without late heading date and one NIL carrying BPH26(t) without spikelet sterility from BC6F2 progeny that showed between 96.3 and 99.8% of the Taichung 65 genetic background through whole-genome survey. In antibiosis testing, the rates of surviving insects and of females with swollen abdomens were lower on the NILs than on Taichung 65, indicating that bph25(t) and Bph26(t) alleles from ADR52 controlled the resistance to BPH. The NILs will serve as useful resources for (1) monitoring BPH virulence and for (2) increasing resistance to BPH.

To broaden the available rice genetic resources, we developed two populations of introgression lines of cultivated rice (Oryza sativa) carrying donor segments from the wild species Oryza glumaepatula and Oryza meridionalis. These lines contain overlapped introgressed donor segments that covered most parts of the genome of the two donors in the same of O. sativa genetic background (ssp. japonica, cv. Taichung 65). The introgression lines were developed through repeated backcrossing with Taichung 65 as a pollen parent and marker assisted selection. O. glumaepatula introgression lines consist of two sets of the lines (with O. glumaepatula and Taichung 65 cytoplasm, respectively): these comprise a total of 69 lines that cover 79.5 to 89.2% of the Oryza glumaepatula genome. The O. meridionalis introgression lines also consist of two sets of the lines (with O. meridionalis and Taichung 65 cytoplasm, respectively): these comprise a total of 78 lines covering the 81.5 to 98.0% of the Oryza meridionalis genome. These introgression lines significantly broaden rice genetic resources, and will facilitate analyses of the genetics of traits specific to the donor species.

We have developed two sets of chromosome segment substitution lines (CSSLs) of cultivated rice (Oryza sativa) carrying donor segments from indica cultivars DV85 and ARC10313. The lines in each set contain chromosomal segments that cover most of the donor genome in a uniform genetic background (ssp. japonica cv. Taichung 65). The starting materials were several recombinant inbred lines derived from the crosses Taichung 65 × DV85 and Taichung 65 × ARC10313. The CSSLs were generated by repeated backcrossing to Taichung 65 (pollen parent), with marker-assisted selection applied at several marker loci. The CSSLs of DV85 (TD-CSSLs) comprise 45 lines that cover 76.6% of the DV85 genome, and the CSSLs of ARC10313 (TA-CSSLs) comprise 44 lines that cover 74.4% of the ARC10313 genome. We investigated the genetic control of days-to-heading in both sets of CSSLs and demonstrated the genetic contribution of several chromosome regions. These CSSLs provide a valuable tool for rice germplasm enhancement, and we expect them to reveal the genetic basis of traits specific to the donor cultivars.

The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is a serious insect pest of cultivated rice (Oryza sativa L.) in temperate East Asia. Six GRH-resistance genes (Grh1, Grh2, Grh3, Grh4, Grh5, and Grh6) and one quantitative trait locus (QTL; qGRH4) have been identified. We selected near-isogenic lines (NILs) carrying Grh1, Grh2, Grh4, Grh5, Grh6, and qGRH4 with the japonica genetic background (Taichung 65 cultivar) by means of marker-assisted selection using new simple sequence repeat markers flanking the GRH-resistance genes and QTL. We also developed three pyramided lines (PYLs; Grh2/Grh6-PYL, Grh4/Grh6-PYL, and Grh5/qGRH4-PYL) using each NIL that carried a GRH-resistance gene or QTL. The NILs, PYLs, and donor parents were evaluated by using an antibiosis test. The resistance of Grh1-NIL and Grh5-NIL did not differ significantly from those of the donor parents, whereas the resistances of Grh2-NIL and Grh6-NIL were significantly lower than those of the donor parents. Grh4-NIL and qGRH4-NIL were highly susceptible. The resistance levels of the pyramided lines for Grh2 and Grh6, Grh4 and Grh6, and Grh5 and qGRH4 demonstrated a gene pyramiding effect that significantly increased their resistance. The developed NILs and PYLs should be useful genetic resources for rice improvement and deployment of the resistance genes.

F1 sterility, one of the most common post-zygotic reproductive barriers, is frequently observed in both interspecific and intraspecific crosses of rice. Elucidating the genetic and cytological mechanisms of F1 pollen sterility is important to exploit genetic resources and to understand the evolutionary dynamics of post-zygotic reproductive isolation in rice. Here, we report two F1 pollen sterility loci, designated S36 and S25, found in an interspecific cross between O. sativa ssp. japonica (Taichung 65) and O. nivara (IRGC105444), and an intraspecific cross between O. sativa ssp. japonica (Asominori) and ssp. indica (IR24). Genetic analyses revealed that both loci are located on distal end of the short arm of chromosome 12 and that allelic interaction at the heterozygous locus caused the sterility of male gametes carrying the japonica alleles in both cases. Comparison of map positions of S36 and S25 suggested that these two loci might be the same locus. Cytological investigation revealed that abnormality of sterile pollen grains caused by S36 occurred mainly at the late bicellular stage after initiation of starch accumulation. This study would provide the better understanding on the genetic nature and the cytological mechanism of F1 pollen sterility in rice.

Deepwater rice (floating rice) can survive under flooded conditions because of their floating ability. We conducted genetic analysis to elucidate the genetic control of floating ability by using an F2 and BC3F2 populations derived from a cross between deepwater and non-deepwater rice varieties. Internode elongation is the most important trait responsible for this adaptation, and is characterized by two factors: timing of the initiation of elongation and the rate of elongation. The position of the lowest elongated internode (LEI) and the rate of internode elongation (RIE) were used to measure floating ability. Two QTLs for LEI were detected on chromosomes 3 (qLEI3) and 12 (qLEI12). For RIE, two QTLs were detected on chromosomes 1 (qRIE1) and 12 (qRIE12). We confirmed the genetic effects and map positions of qLEI3, qLEI12 and qRIE12 by using BC3F2 populations. Characterization of near-isogenic lines of qLEI3, qLEI12 and qRIE12 revealed that the LEI and RIE are at least partly controlled by different genetic pathways. Observation of near-isogenic lines suggested that the introgressed segment of qLEI12 and qRIE12 of chromosome 12 affected the deepwater-responsive elongation.

To gain a comprehensive understanding of the genetic regulation of male gametogenesis, we screened and identified mutants for abnormal pollen development in rice (Oryza sativa L.). We gamma-irradiated spikelets of a japonica rice cultivar, Taichung 65, just before and after anthesis. Among 1,000 M2 lines, 24 lines were identified as mutants for pollen sterility. Progeny tests in M3 demonstrated that pollen sterility in 12 lines was controlled by single recessive genes, designated sps1–sps12 (sporophytic pollen sterility). In the remaining 12 lines, pollen semi-sterility was under the control of gametophytic genes, designated gps1–gps12 (gametophytic pollen sterility). Linkage analysis revealed that sps6, sps9, and sps12 were mapped on chromosomes 3, 9, and 7, respectively.

Rice bran oil is a byproduct of the milling of rice (Oryza sativa L.). It offers various health benefits and has a beneficial fatty acid composition. To increase the amount of rice bran as a sink for triacylglycerol (TAG), we developed and characterized new breeding materials with giant embryos. To induce mutants, we treated fertilized egg cells of the high-yielding cultivar ‘Mizuhochikara’ with N-methyl-N-nitrosourea (MNU). By screening M2 seeds, we isolated four giant embryo mutant lines. Genetic analysis revealed that the causative loci in lines MGE12 and MGE13 were allelic to giant embryo (ge) on chromosome 7, and had base changes in the causal gene Os07g0603700. On the other hand, the causative loci in lines MGE8 and MGE14 were not allelic to ge, and both were newly mapped on chromosome 3. The TAG contents of all four mutant lines increased relative to their wild type, ‘Mizuhochikara’. MGE13 was agronomically similar to ‘Mizuhochikara’ and would be useful for breeding for improved oil content.

Chromosome substitution series are useful tools for precise mapping of quantitative trait loci (QTLs) and for the evaluation of gene action as a single factor or in the case of gene interactions. In this study, we produced reciprocal sets of chromosome segment substitution lines (CSSLs) between Asominori (Japonica) and IR24 (Indica) in cultivated rice, Oryza sativa L. A set of chromosome segment substitution lines carries a single or a few donor chromosome segments overlapping with neighboring segments in a uniform genetic background. Reciprocal sets were developed by repeated backcrossing to respective recurrent parents and DNA marker-assisted selections. The sets consisted of a series of 70 lines of Asominori CSSLs with IR24 genetic background (designated as ‘IAS’) and a series of 91 lines of IR24 CSSLs with Asominori genetic background (designated as ‘AIS’). These substitution series represented most of the genome of the donor parents on the alien genetic backgrounds. Each of the substitution lines was nearly isogenic to the recurrent parent: the average proportions of recurrent parent’s genome were 93 % and 91 % in the IAS and AIS, respectively. Consistent reciprocal QTL effects for heading date and grain size were observed in both IAS and AIS. The reciprocal substitution set appeared to be an efficient and systematic method for QTL precision analysis.

The brown planthopper(BPH), Nilaparvata lugens Stal, is a serious pest of rice(Oryza sativa L.) in Asia. The rice ovicidal response to BPH is characterized by the formation of watery lesions which result in the death of the eggs. It is one of the factors affecting the suppression of the multiplication of BPH. To detect quantitative trait loci(QTLs) for this ovicidal response, a set of 71 rice recombinant inbred lines(F11) derived from a cross between a Japonica variety Asominori with ovicidal response and an Indica variety IR24 without ovicidal response were phenotyped for grade of watery lesions(GWL) and egg mortality(EM) of BPH. GWL and EM showed a significant positive correlation(P

GRAIN SIZE 3 (GS3) is a cloned gene that is related to seed length. Here we report the discovery of new deletion alleles at the GS3 locus, each of which confer short seed. We selected ten short seeded cultivars from a collection of 282 diverse cultivars. Sequence analysis across the GS3 gene in these ten cultivars identified three novel alleles and a known allele that contain several independent deletion(s) in the fifth exon of GS. These independent deletion variants each resulted in a frameshift mutation that caused a premature stop codon, and they were functionally similar to one another. Each coded for a truncated gene product that behaved as an incomplete dominant allele and conferred a short seeded phenotype. Haplotype analysis of these sequence variants indicated that two of the variants were of japonica origin, and two were from indica. Transformation experiments demonstrated that one of the deletion alleles of GS3 decrease the cell number in the upper epidermis of the glume, resulting in a significant reduction in seed length. The multiple and independent origins of these short seeded alleles indicate that farmers and early breeders imposed artificial selection favoring short seeds.