Quantitative trait locus analyses of ozone-induced grain yield reduction in rice

Reduction of grain yield (total seed weight) by ozone in rice (Oryza sativa L.) is believed to be caused by ozone-induced reduction of photosynthetic activity followed by growth inhibition. Here, japonica rice cultivar Sasanishiki and indica rice cultivar Habataki showed different responses to ozone. When exposed to ozone, the leaves of Habataki exhibited no critical damage, whereas those of Sasanishiki developed lesions. Conversely, ozone exposure reduced total seed weight by 19% in Habataki, but not significantly in Sasanishiki. Chronic ozone exposure also significantly decreased culm length, number of primary rachis branch, and number of spikelets per panicle in Habataki. QTL analysis in Sasanishiki/Habataki chromosome segment substitution lines identified a single locus associated with the yield loss caused by ozone on chromosome 6 of Habataki close to marker RM3430 (107.6 cM). A QTL for reduction of primary rachis branch number and total spikelet number was found in the same position. These results indicate that a QTL on chromosome 6 has an important role in ozone-induced yield loss, and is also involved in primary rachis branch formation and total spikelet number in ozone-exposed rice.     

粳稻穗部结构性状的QTL分析

利用温带粳稻‘沈农265’和‘丽江新团黑谷’构建的重组自交系群体,在沈阳和哈尔滨两地对15个穗部结构性状进行了QTL分析。共检测到61个相关QTL,其中沈阳检测到的38个QTL在第1、4、6、11和12号染色体上形成了sir-QTL簇;而在哈尔滨检测到的31个QTL也在第3、9和10号染色体上形成了QTL簇。仅有8个QTL是在两地同时被检测到的,分别是控制一次枝梗数#＇．jqPBN4、控制穗长的qPL6和qPL9、控制一次枝梗实粒数的qGNPB4、控制一次枝梗颖花数的qTSNPBJ、控制结实率的qPSSIO、以及控制着粒密度qSD3和qSD9。其中,qPBN4（最高表型贡献率43．2％）、qPL9（最高表型贡献率63．2％）、qGNPB4（最高表型贡献率30．9％）和qSD9（最高表型贡献率42．9％）是主效QTL。通过进一步的分析发现控制穗长qPL9和控制着粒密度qSD94于DEPl所在区间。同时控制一次枝梗数和一次枝梗实粒数的位于第4号染色体长臂端的穗部结构主效QTL,qPBN4qGNPB4极富研究与应用价值。     

Mapping quantitative trait loci for heat tolerance at the booting stage using chromosomal segment substitution lines in rice

High temperature stress is a major obstacle in rice productivity. Considerable progress has been made on studying heat tolerance (HT) at different stages. However, the genetic basis of HT at the booting stage is poorly understood. In this study, we analyzed the morphological features of a heat-sensitive japonica cultivar Sasanishiki under natural high temperature stress at the booting stage. The anthers became smaller and the number, and fertility, of pollen grains were decreased significantly. As a result, there was a dramatic reduction in spikelet fertility. In contrast, the indica cultivar Habataki showed high HT and normal spikelet fertility under high temperature stress. Additonally, a set of chromosome segment substitution lines, derived from Sasanishiki and Habataki, were evaluated for HT related quantitative trait loci (QTLs) across two environments in the natural field. A total of 12 QTLs associated with HT were detected, of which, 5 were identified in two environments, and 7 in one environment. Furthermore, one of the major-effect QTLs (qHTB3-3) detected on the long arm of chromosome 3, was confirmed using overlapping substituted lines. qHTB3-3 was finally mapped between the two markers RM3525 and 3-M95, approximately 2.8 Mb apart. These findings and further gene cloning of qHTB3-3 will help us better understand the molecular control of HT in rice, and may contribute to the development of high HT rice varieties.     

Development of Chromosome Segment Substitution Lines Derived from Backcross between Two Sequenced Rice Cultivars, <Emphasis Type="Italic">Indica</Emphasis> Recipient 93-11 and <Emphasis Type="Italic">Japonica</Emphasis> Donor Nipponbare

Chromosome segment substitution lines (CSSLs) are powerful tools for detecting and precisely mapping quantitative trait loci (QTLs) and evaluating gene action as a single factor. In this study, 103 CSSLs were produced using two sequenced rice cultivars: 93-11, an elite restorer indica cultivar as recipient, and Nipponbare, a japonica cultivar, as donor. Each CSSL carried a single chromosome substituted segment. The total length of the substituted segments in the CSSLs was 2,590.6 cM, which was 1.7 times of the rice genome. To evaluate the potential application of these CSSLs for QTL detection, phenotypic variations of seed shattering, grain length and grain width in 10 CSSLs were observed. Two QTLs for seed shattering and three for grain length and grain width were identified and mapped on rice chromosomes. The results demonstrate that CSSLs are excellent genetic materials for dissecting complex traits into a set of monogenic loci. These CSSLs are of great potential value for QTL mapping and plant marker-assisted breeding (MAB).     

A gene controlling the number of primary rachis branches also controls the vascular bundle formation and hence is responsible to increase the harvest index and grain yield in rice

The quantitative trait locus controlling the number of primary rachis branches (PRBs) in rice was identified using backcrossed inbred lines of Sasanishiki/Habataki//Sasanishiki///Sasanishiki. The resultant gene was ABERRANT PANICLE ORGANIZATION 1 (APO1). Habataki-genotype segregated reciprocal recombinant lines for the APO1 locus increased both the number of PRB (12–13%) and the number of grains per panicle (9–12%), which increased the grain yield per plant (5–7%). Further recombination dividing this region revealed that different alleles regulated the number of PRB and the number of grains per panicle. The PRB1 allele, which includes the APO1 open reading frame (ORF) and the proximal promoter region, controlled only the number of PRB but not the number of grains per panicle. In contrast, the HI1 allele, which includes only the distal promoter region, increased the grain yield and harvest index in Habataki-genotype plants, nevertheless, the ORF expressed was Sasanishiki type. It also increased the number of large vascular bundles in the peduncle. APO1 expression occurred not only in developing panicles but also in the developing vascular bundle systems. In addition, Habataki plants displayed increased APO1 expression in comparison to Sasanishiki plants. It suggests that APO1 enhances the formation of vascular bundle systems which, consequently, promote carbohydrate translocation to panicles. The HI1 allele is suggested to regulate the amount of APO1 expression, and thereby control the development of vascular bundle systems. These findings may be useful to improve grain yield as well as quality through the improvement of translocation efficiency.     

Temperature-dependent QTLs in <Emphasis Type="Italic">indica</Emphasis> alleles for improving grain quality in rice: increased prominence of QTLs responsible for reduced chalkiness under high-temperature conditions

Quantitative trait loci (QTLs) for the apparent quality of brown rice under high temperatures during ripening were analyzed using chromosomal segment substitution lines. Segments from the indica cultivar Habataki were substituted into a japonica cultivar with a Sasanishiki background. We found the following two QTLs for increasing grain quality in the Habataki allele on chromosome 3: (1) qTW3-2, located near the marker RM14702, decreased the percentage of total white immature (TWI) grains, and (2) qRG3-2, located near RM3766, increased the percentage of regular grains. The effects of these two QTLs were more obvious under high-temperature ripening conditions; hence, these loci are considered QTLs not only for reducing TWI grains but also for increasing high-temperature tolerance. Additionally, we found two QTLs, i.e., qTW3-1 and qRG3-1, responsible for reduced grain quality near RM14314 on chromosome 3. Although the QTL for narrow grains in the Habataki allele qNG3 was genetically linked to qTW3-2, the effect was only slightly significant, and the length/width ratio of qNG3-carrying grains was within the range observed in widely grown japonica cultivars. Incorporating the Habataki region, including qRG3-2 and qTW3-2 but not qTW3-1 and qRG3-1, in addition to previously reported grain quality QTLs in breeding japonica cultivars will improve high-temperature tolerance and grain quality.     

The<Emphasis Type="Italic"> Rxo1</Emphasis>/<Emphasis Type="Italic">Rba1</Emphasis> locus of maize controls resistance reactions to pathogenic and non-host bacteria

The genetic mechanism underlying six palatability properties of cooked rice and three physico-chemical traits was dissected in 66 BC₃F₂ chromosome segment substitution lines (CSSLs), using a complete linkage map in three successive years. The CSSLs showed transgressive segregation for all traits studied. Significant correlation was detected among most palatability traits. A total of 25 QTLs for the nine traits were identified on nine chromosomes, and many QTLs affecting different quality traits were mapped in the same regions. Six QTLs—qLT-8 for luster, qTD-6 and qTD-8 for tenderness, qIVOE-6 and qIVOE-8 for integrated value of organoleptic evaluation, and qAC-8 for amylose content—were repeatedly detected across the 3 years. Phenotypic values were significantly different between the recurrent parent, cultivar Asominori, and the CSSLs harboring any of the six QTL alleles across the three environments, indicating that these six QTLs were non-environment-specific and could be used for marker-assisted selection in rice quality improvement.     

Mapping and characterization of seed dormancy QTLs using chromosome segment substitution lines in rice

Seed dormancy—the temporary failure of a viable seed to germinate under favorable conditions—is a complex characteristic influenced by many genes and environmental factors. To detect the genetic factors associated with seed dormancy in rice, we conducted a QTL analysis using chromosome segment substitution lines (CSSLs) derived from a cross between Nona Bokra (strong dormancy) and Koshihikari (weak dormancy). Comparison of the levels of seed dormancy of the CSSLs and their recurrent parent Koshihikari revealed that two chromosomal regions—on the short arms of chromosomes 1 and 6—were involved in the variation in seed dormancy. Further genetic analyses using an F₂ population derived from crosses between the CSSLs and Koshihikari confirmed the allelic differences and the chromosomal locations of three putative QTLs: Sdr6 on chromosome 1 and Sdr9 and Sdr10 on chromosome 6. The Nona Bokra alleles of the three QTLs were associated with decreased germination rate. We discuss the physiological features of the CSSLs and speculate on the possible mechanisms of dormancy in light of the newly detected QTLs.     

A Pyramid Breeding of Eight Grain-yield Related Quantitative Trait Loci Based on Marker-assistant and Phenotype Selection in Rice （Oryza sativa L.）

1000-Grain weight and spikelet number per panicle are two important components for rice grain yield.In our previous study,eight quantitative trait loci(QTLs)conferring spikelet number per panicle and 1000-grain weight were mapped through sequencing-based genotyping of 150 rice recombinant inbred lines(RILs).In this study,we validated the effects of four QTLs from Nipponbare using chromosome segment substitution lines(CSSLs),and pyramided eight grain yield related QTLs.The new lines containing the eight QTLs with positive effects showed increased panicle and spikelet size as compared with the parent variety 93-11.We further proposed a novel pyramid breeding scheme based on marker-assistant and phenotype selection(MAPS).This scheme allowed pyramiding of as many as 24 QTLs at a single hybridization without massive cross work.This study provided insights into the molecular basis of rice grain yield for direct wealth for high-yielding rice breeding.     

QTL Analysis of Aluminum Resistance in Rice (Oryza sativa L.)

Aluminum (Al) toxicity is considered as one of the primary causes of low-rice productivity in acid soils. In the present study, quantitative trait loci (QTLs) controlling Al resistance based on relative root elongation (RRE) were dissected using a complete linkage map and a recombinant inbred lines (RILs) derived from a cross of Al-tolerant japonica cultivar Asominori (Oryza sativa L.) and Al-sensitive indica cultivar IR24 (O. sativa L.). A total of three QTLs (qRRE-1, qRRE-9, and qRRE-11) were detected on chromosomes 1, 9, and 11 with LOD score ranging from 2.64 to 3.60 and the phenotypic variance explained from 13.5 to 17.7%. The Asominori alleles were all associated with Al resistance at all the three QTLs. The existence of these QTLs was confirmed using Asominori chromosome segment substitution lines (CSSLs) in IR24 genetic background (IAS). By QTL comparative analysis, the two QTLs (qRRE-1and qRRE-9) on chromosomes 1 and 9 appeared to be consistent among different rice populations while qRRE-11 was newly detected and syntenic with a major Al resistance gene on chromosome 10 of maize. This region may provide an important case for isolating genes responsible for different mechanisms of Al resistance among different cereals. These results also provide the possibilities of enhancing Al resistance in rice breeding program by marker-assisted selection (MAS) and pyramiding QTLs.     

Indian rice “Kasalath” contains genes that improve traits of Japanese premium rice “Koshihikari”

Rice (Oryza sativa L.) chromosome segment substitution lines (CSSLs), in which chromosomal segments of the Indian landrace “Kasalath” replace the corresponding endogenous segments in the genome of the Japanese premium rice “Koshihikari”, are available and together cover the entire genome. Chromosome regions affecting a trait (CRATs) can be identified by comparison of phenotypes with genotypes of CSSLs. We detected 99 CRATs for 15 agronomic or morphological traits. “Kasalath” had positively acting alleles in 53 CRATs. Its CRATs increased panicle number per plant by up to 23.3%, grain number per panicle by up to 30.8%, and total grain number by up to 15.1%, relative to “Koshihikari”. CRATs were identified for grain size (grain thickness and width), with positive effects of about 5.0%. A CRAT on chromosome 8 almost doubled the weight of roots in uppermost soil layers compared to “Koshihikari”. Additionally, “Kasalath” possessed CRATs for higher lodging resistance (reduction in plant height and increase in stem diameter). In some cases, multiple CRATs were detected in the same chromosome regions. Therefore, CSSLs with these chromosome segments might be useful breeding materials for the simultaneous improvement of multiple traits. Five CRATs, one for plant height on chromosome 1, one for stem diameter on chromosome 8, and three for heading date on chromosomes 6, 7, and 8 overlapped with the corresponding QTLs that already had been mapped with back-crossed inbred lines of “Nipponbare” and “Kasalath”. In both “Koshihikari” CRATs and “Nipponbare” QTLs, “Kasalath” had similar effects. Both Y. Madoka and T. Kashiwagi have contributed equally to this article.     

Chromosomal regions with quantitative trait loci controlling cadmium concentration in brown rice (Oryza sativa)

A novel mapping population consisting of 39 chromosome segment substitution lines (CSSLs) was used to locate the putative quantitative trait loci (QTLs) for cadmium (Cd) concentration in brown rice (Oryza sativa). The mapping population carried a single chromosome segment of 'Kasalath' (indica) in each line overlapping with neighbouring segments in a 'Koshihikari' (japonica) genetic background. The parents and CSSLs were grown in pots filled with Cd-polluted soil until grain filling. The brown rice of three of the 39 CSSLs had significantly lower Cd concentrations than that of Koshihikari, and the brown rice of a further three had significantly higher concentrations. On the basis of graphical genotypes of CSSLs, putative QTLs controlling the Cd concentration in brown rice were detected on chromosomes 3, 6 and 8. Each of the CSSLs was nearly isogenic to Koshihikari, which is the most popular rice cultivar in Japan: they carried > 90% of the Koshihikari genetic background. Therefore, the development of a new Koshihikari with less Cd concentration in brown rice would be feasible in the near future.     

Genetic analysis of root elongation induced by phosphorus deficiency in rice (Oryza sativa L.): fine QTL mapping and multivariate analysis of related traits

Root elongation induced by phosphorus deficiency has been reported as one of the adaptive mechanisms in plants. Genetic differences were found in rice for the root elongation under phosphorus deficiency (REP), for which a distinct quantitative trait locus (QTL) was detected on the long arm of chromosome 6. Subsequently, the effect and position of the QTL, designated as qREP-6, were confirmed using chromosome segment substitution lines (CSSLs), in which the background of a japonica cultivar, ‘Nipponbare’ with non-REP, was partially substituted by chromosomal segments from an indica cultivar, ‘Kasalath’ with remarkable REP. Out of 54 CSSLs, two lines (CSSL28 and CSSL29) that retain a common ‘Kasalath’-derived segment on the long arm of chromosome 6 showed a significantly high REP. The high REP lines also showed high adaptabilities such as enhanced tillering ability and shoot phosphorus content. Accordingly, conditional dependencies between the related traits were assessed using a graphical Gaussian model (GGM). Direct interactions between REP and root length, and between root length and tiller number were detected under P deficiency in CSSLs. Furthermore, qREP-6 for REP and qTNP-6 for tiller number under P deficiency were fine-mapped with an F₂ population of a cross between Nipponbare and CSSL29. A region containing qREP-6 accounted for more than half of the phenotypic variance, the most plausible interval of which contained 37 candidate genes. The result provides a foundation for cloning of the qREP-6 gene which will be applicable to study P deficiency-dependent response and to improve rice’s adaptability to P deficiency stress. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

水稻叶绿素含量动态QTL分析

为剖析水稻不同生育时期叶绿素含量的变化动态及遗传机制,以‘Sasanishiki’（粳稻）、‘Habataki’（籼稻）及其杂交衍生的85个回交重组自交系（BILs）群体为材料,对控制水稻叶片叶绿素含量的数量性状基因位点（QTL）变化动态进行了分析。共检测到39个QTL,包括26个非条件QTL和13个条件QTL,分布在除第7和第11号染色体以外的10条染色体上,平均每个时期检测到3．25个非条件QTL。其中生育前期和生育中后期检测到的QTL位点较少,仅为1—3个;在生育中期（盛期）检测到的QTL位点相对较多,一般为4～5个。在生育期的中期和后期均能在第1和2号染色体上检测到控制叶绿素含量的QTL,并且这些QTL位点在1和2号染色体上呈现聚集现象。本研究同时发现了一些新的QTL位点,这些QTL将有助于我们更全面地了解叶绿素在不同发育时期的遗传基础。     

A major locus qS12, located in a duplicated segment of chromosome 12, causes spikelet sterility in an indica-japonica rice hybrid

Chromosome segment duplications are integral in genome evolution by providing a source for the origin of new genes. In the rice genome, besides an ancient polyploidy event known in the rice common ancestor, it had been identified that there was a special segmental duplication involving chromosomes 11 and 12, but the biological role of this duplication remains unknown. In this study, by using a set of chromosome segment substitution lines (CSSLs) and near isogenic lines (NILs) derived from the indica cultivar 9311 and japonica cultivar Nipponbare, a major QTL (qS12) resulting in hybrid male sterility was mapped within ~400 kb region adjacent to the special duplicated segment on the short arm of chromosome 12. Compared to the japonica cultivar Nipponbare, the two sides of the qS12 candidate region were inverted in the indica cultivar 9311. Among 47 of the 111 rice genotypes evaluated by molecular markers, the inverted sides were detected, and found completely homologous to indica cultivar 9311. These results suggested that the two inverted sides protect the sequence in the qS12 regions from recombination. On the short-arm of chromosome 12, two QTLs S-e and S25, in addition to qS12, were previously detected as a distinct segregation distortion and pollen semi-sterility loci. We propose these three hybrid sterility loci are the same locus, and the duplicated segment on chromosome 12 may play a prominent role in diversification, i.e., sub-speciation of cultivated rice.     

Fine mapping of a yield-enhancing QTL cluster associated with transgressive variation in an <Emphasis Type="Italic">Oryza sativa </Emphasis>× <Emphasis Type="Italic">O. rufipogon</Emphasis> cross

A high-resolution physical map targeting a cluster of yield-related QTLs on the long arm of rice chromosome 9 has been constructed across a 37.4 kb region containing seven predicted genes. Using a series of BC₃F₄ nearly isogenic lines (NILs) derived from a cross between the Korean japonica cultivar Hwaseongbyeo and Oryza rufipogon (IRGC 105491), a total of seven QTLs for 1,000-grain weight, spikelets per panicle, grains per panicle, panicle length, spikelet density, heading date and plant height were identified in the cluster (P ≤ 0.0001). All seven QTLs were additive, and alleles from the low-yielding O. rufipogon parent were beneficial in the Hwaseongbyeo background. Yield trials with BC₃F₄ NILs showed that lines containing a homozygous O. rufipogon introgression in the target region out-yielded sibling NILs containing Hwaseongbyeo DNA by 14.2–17.7%, and out-yielded the Hwaseongbyeo parent by 16.2–23.7%. While higher yielding plants containing the O. rufipogon introgression were also taller and later than controls, the fact that all seven of the QTLs were co-localized in the same 37.4 kb interval suggests the possibility that a single, pleiotropic gene acting as a major regulator of plant development may control this suite of agronomically important plant phenotypes. Xiaobo Xie and Fengxue Jin have contributed equally to this study.     

Role of the iron transporter OsNRAMP1 in cadmium uptake and accumulation in rice

The heavy metal cadmium (Cd) is toxic to humans, and its accumulation in rice grains is a major agricultural problem. Rice has seven putative metal transporter NRAMP genes, but microarray analysis showed that only OsNRAMP1 is highly up-regulated by iron (Fe) deficiency. OsNRAMP1 localized to the plasma membrane and transported Cd as well as Fe. OsNRAMP1 expression was observed mainly in roots and was higher in the roots of a high-Cd-accumulating cultivar (Habataki) than in those of a low-Cd-accumulating cultivar (Sasanishiki). The amino acid sequence of OsNRAMP1 in the Sasanishiki and Habataki cultivars was found to be 100% identical. These results suggest that OsNRAMP1 participates in cellular Cd uptake and that the differences observed in Cd accumulation among cultivars are because of differences in OsNRAMP1 expression levels in roots.     

Genome‐wide expression quantitative trait locus studies facilitate isolation of causal genes controlling panicle structure

The morphology of rice (Oryza sativa L.) panicles is an important determinant of grain yield, and elucidation of the genetic control of panicle structure is very important for fulfilling the demand for high yield in breeding programs. In a quantitative trait locus (QTL) study using 82 backcross inbred lines (BILs) derived from Koshihikari and Habataki, 68 QTLs for 25 panicle morphological traits were identified. Gene expression profiling from inflorescence meristems of BILs was obtained. A combination of phenotypic QTL (pQTL) and expression QTL (eQTL) analysis revealed co‐localization between pQTLs and eQTLs, consistent with significant correlations between phenotypic traits and gene expression levels. By combining pQTL and eQTL data, two genes were identified as controlling panicle structure: OsMADS18 modulates the average length of the primary rachis and OsFTL1 has pleiotropic effects on the total number of secondary rachides, number of grains per panicle, plant height and the length of flag leaves. Phenotypes were confirmed in RNA interference knocked‐down plants and overexpressor lines. The combination of pQTL and eQTL analysis could facilitate identification of genes involved in rice panicle formation.     

Identification of quantitative trait loci controlling rice mature seed culturability using chromosomal segment substitution lines

The genetic transformation efficiency of a rice variety is largely determined by its tissue culturability. Establishment of a highly efficient tissue-culture system has greatly accelerated the wide spread application of transgenic japonica varieties. However, such process for indica rice was hampered because this type of variety is recalcitrant to in vitro culture. This study aimed to map the quantitative trait loci (QTLs) for mature seed culturability using a chromosomal segment substitution lines (CSSL) population derived from a cross between an indica variety “Zhenshan 97B” and a japonica variety “Nipponbare”. The CSSLs consist of 139 lines each containing a single or a few introgression segments, and together covering the whole “Nipponbare” genome. Every CSSL was tested by culturing on the two medium systems developed for the respective indica and japonica parental varieties. The performance of culturability was evaluated by four indices: frequency of callus induction (CIF), callus subculture capability (CSC), frequency of plant regeneration (PRF) and the mean plantlet number per regenerated callus (MNR). All four traits displayed continuous variation among the CSSLs. With the culture system for japonica rice, three CIF QTLs, three CSC QTLs, three PRF QTLs and three MNR QTLs were detected. With the culture system for indica variety, six CIF QTLs, two CSC QTLs, three PRF QTLs and six MNR QTLs were identified, and these QTLs distributed on nine rice chromosomes. Two QTLs of CIF and two QTLs of MNR were detected in both the japonica and indica rice culture system. The correlation coefficients of all the four traits varied depending on the culture systems. These results provide the possibilities of enhancing the culturability of indica rice by marker-assisted breeding with those desirable alleles from the japonica. Lina Zhao and Hongju Zhou have contributed equally to this work.