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.     

Identification of qRBS1, a QTL involved in resistance to bacterial seedling rot in rice

Bacterial seedling rot (BSR), a destructive disease of rice (Oryza sativa L.), is caused by the bacterial pathogen Burkholderia glumae. To identify QTLs for resistance to BSR, we conducted a QTL analysis using chromosome segment substitution lines (CSSLs) derived from a cross between Nona Bokra (resistant) and Koshihikari (susceptible). Comparison of the levels of BSR in the CSSLs and their recurrent parent, Koshihikari, revealed that a region on chromosome 10 was associated with resistance. Further genetic analyses using an F₅ population derived from a cross between a resistant CSSL and Koshihikari confirmed that a QTL for BSR resistance was located on the short arm of chromosome 10. The Nona Bokra allele was associated with resistance to BSR. Substitution mapping in the Koshihikari genetic background demonstrated that the QTL, here designated as qRBS1 (quantitative trait locus for RESISTANCE TO BACTERIAL SEEDLING ROT 1), was located in a 393-kb interval (based on the Nipponbare reference genome sequence) defined by simple sequence repeat markers RM24930 and RM24944.     

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).     

Development and high-throughput genotyping of substitution lines carring the chromosome segments of indica 9311 in the background of japonica Nipponbare

Chromosome segment substitution lines (CSSLs) are useful for the precise mapping of quartitative trait loci (QTLs) and dissection of the genetic basis of complex traits.In this study,two whole-genome sequenced rice cultivars,the japonica Nipponbare and indica 9311 were used as recipient and dtonor,respectively.A population with 57 CSSLs was developed after crossing and back-crossing assisted by mo lecular rnarkers,and genotypes were identified using a high-throughput resequencing strategy,Detailed graphical genotypes of 38 lines were constructed based on resequencing data.These CSSLs had a total of 95 substituted segments derived from indica 9311,with an average of about 2.5 segments pet CSSL and eight segments per chromosome,and covered about 87.4％ of the rice whole genome.A multiple linear regression QTL analysis mapped four QTLs for 1000-grain weight.The largest-effect QTL was located in a region on chromosome 5 that contained a cloned major QTL GW5/qSW5 for grain size in rice.These CSSLs with a background of Nipponbare may provide powerful tools for future whole-genome discovery and functional study of essential genes/QTLs in rice,and offer ideal materials and foundations for japonica breeding.     

A Chromosome Segment Substitution Library of Weedy Rice for Genetic Dissection of Complex Agronomic and Domestication Traits

Chromosome segment substitution lines (CSSLs) are a powerful alternative for locating quantitative trait loci (QTL), analyzing gene interactions, and providing starting materials for map-based cloning projects. We report the development and characterization of a CSSL library of a U.S. weedy rice accession ‘PSRR-1’ with genome-wide coverage in an adapted rice cultivar ‘Bengal’ background. The majority of the CSSLs carried a single defined weedy rice segment with an average introgression segment of 2.8 % of the donor genome. QTL mapping results for several agronomic and domestication traits from the CSSL population were compared with those obtained from two recombinant inbred line (RIL) populations involving the same weedy rice accession. There was congruence of major effect QTLs between both types of populations, but new and additional QTLs were detected in the CSSL population. Although, three major effect QTLs for plant height were detected on chromosomes 1, 4, and 8 in the CSSL population, the latter two escaped detection in both RIL populations. Since this was observed for many traits, epistasis may play a major role for the phenotypic variation observed in weedy rice. High levels of shattering and seed dormancy in weedy rice might result from an accumulation of many small effect QTLs. Several CSSLs with desirable agronomic traits (e.g. longer panicles, longer grains, and higher seed weight) identified in this study could be useful for rice breeding. Since weedy rice is a reservoir of genes for many weedy and agronomic attributes, the CSSL library will serve as a valuable resource to discover latent genetic diversity for improving crop productivity and understanding the plant domestication process through cloning and characterization of the underlying genes.     

Time-related identification of chromosome regions affecting plant elongation in rice (Oryza sativa L.).

We investigated the time-related changes of Chromosome Regions that Affect Traits (CRATs) for elongation rate in rice (Oryza sativa L.) using chromosome segment substitution lines (CSSLs) carrying a single chromosome segment of the cultivar Kasalath (indica) in a Koshihikari (japonica) genetic background. The growth period of rice was partitioned into eight stages (each lasting 5-7days) from 18days after transplanting, and the elongation rate was determined as the increase of total plant height per time at each growth stage. CRATs for plant elongation rate were determined based on graphical genotype data of CSSLs that showed a significantly higher or lower elongation rate than Koshihikari. In total, 23 CRATs for plant elongation rate were detected, and different CRATs acted at different growth stages. Fifteen CRATs increased the elongation rate through Kasalath alleles, and eight increased it through Koshihikari alleles. These results suggest that plant height in rice is regulated in a stage-specific manner by a variety of genetic mechanisms that control plant elongation rate. Kasalath alleles of PE1-9 increased the elongation rate at an early growth stage (18-25days after transplanting), while Koshihikari alleles of PE8-3 decreased the elongation rate at a late growth stage (68-74days after transplanting). In a line that contained both of these CRATs, the elongation rate at the early growth stage was increased without affecting plant height at harvesting. We conclude that stage-specific optimization of plant height in rice may be achieved by combining CRATs that control plant elongation at specific stages.     

Genetic effects of major QTLs controlling low-temperature germinability in different genetic backgrounds in rice (Oryza sativa L.)

The effects of QTLs are demonstrated basically within the population used in the original QTL analysis as the difference between the alleles of the parental varieties. For the efficient use of QTLs in breeding programs, it is necessary to assess whether the QTL exhibits its genetic effect when it is introgressed into different genetic backgrounds. Extensive studies of tolerance to low temperature at the seed germination stage (called low-temperature germinability) in rice revealed that 2 major QTLs on chromosomes 3 and 11, qLTG3-1 and qLTG11, have large effects. This study assessed the effects of these 2 QTLs from an aus variety, Kasalath, in different genetic backgrounds of 3 japonica varieties, Hoshinoyume, Hayamasari, and Koshihikari. Backcrossed progenies and chromosome segment substitution lines showed that both qLTG3-1 and qLTG11 were effective in the Hoshinoyume and Koshihikari backgrounds, while only qLTG3-1 was effective in the Hayamasari background. The results in this study demonstrated that these 2 QTLs are useful for the improvement of low-temperature germinability in rice breeding programs. The results also indicated that low-temperature germinability in rice is governed by an epistatic interaction of qLTG11.     

Identification of Quantitative Trait Loci for Rice Quality in a Population of Chromosome Segment Substitution Lines

The demand for high quality rice represents a major issue in rice production. The primary components of rice grain quality include appearance, eating, cooking, physico-chemical, milling and nutritional qualities. Most of these traits are complex and controlled by quantitative trait loci (QTLs), so the genetic characterization of these traits is more difficult than that of traits controlled by a single gene. The detection and genetic identification of QTLs can provide insights into the genetic mechanisms underlying quality traits. Chromosome segment substitution lines (CSSLs) are effective tools used in mapping QTLs. In this study, we constructed 154 CSSLs from backcross progeny (BC3F2) derived from a cross between 'Koshihikari' (an Oryza sativa L. Ssp. Japonica variety) as the recurrent parent and 'Nona Bokra' (an O. Sativa L. Ssp. Indica variety) as the donor parent. In this process, we carried out marker-assisted selection by using 102 cleaved amplified polymorphic sequence and simple sequence repeat markers covering most of the rice genome. Finally, this set of CSSLs was used to identify QTLs for rice quality traits. Ten QTLs for rice appearance quality traits were detected and eight QTLs concerned physico-chemical traits. These results supply the foundation for further genetic studies and breeding for the improvement of grain quality.     

Development of nutritious rice with high zinc/selenium and low cadmium in grains through QTL pyramiding

Enriching zinc (Zn) and selenium (Se) levels, while reducing cadmium (Cd) concentration in rice grains is of great benefit for human diet and health. Large natural variations in grain Zn, Se, and Cd concentrations in different rice accessions enable Zn/Se‐biofortification and Cd‐minimization through molecular breeding. Here, we report the development of new elite varieties by pyramiding major quantitative trait loci (QTLs) that significantly contribute to high Zn/Se and low Cd accumulation in grains. A chromosome segment substitution line CSSL^GCC7 with the PA64s‐derived GCC7 allele in the 93‐11 background, exhibited steadily higher Mn and lower Cd concentrations in grains than those of 93‐11. This elite chromosome segment substitution line (CSSL) was used as the core breeding material to cross with CSSLs harboring other major QTLs for essential mineral elements, especially CSSL^GZC6 for grain Zn concentration and CSSL^GSC5 for grain Se concentration. The CSSL^GCC7+GZC6 and CSSL^GCC7+GSC5 exhibited lower Cd concentration with higher Zn and Se concentrations in grains, respectively. Our study thus provides elite materials for rice breeding targeting high Zn/Se and low Cd concentrations in grains.     

利用粳稻染色体片段置换系群体检测水稻抗亚铁毒胁迫有关性状QTL

潜育性水稻田广泛分布于中国、斯里兰卡、印度、印度尼西亚、塞拉里昂、利比亚、尼日利亚、哥伦比亚和菲律宾等国,其中我国南方稻区就有近700万公顷低产潜育性水稻田。该类水稻田还原性强,矿质营养失调,尤以Fe^2 过量积累,对水稻生长发育产生不良的逆境胁迫作用。培育抗亚铁毒的水稻品种是简便、经济有效地提高稻谷产量的重要途径之一。该文利用由粳稻品种Asominori与籼稻品种IR24杂交衍生的Asominori染色体片段置换系(Chromosome Segment Substitution Lines,CSSLs)群体为材料,检测与抗亚铁毒胁迫有关性状QTL。共检测到与抗亚铁毒胁迫有关性状QTL14个,各QTL的LOD值为2．72～6．63。其中检测到与抗亚铁毒胁迫直接有关的性状叶片棕色斑点指数QTL3个,分别位于第3、9、11染色体C515～XNpb279、R2638～C1263和G1465～C950之间,对应的贡献率分别为16．45％、11．16％和28．02％;与其他已发表的定位结果比较发现,位于第三染色体C515～XNpb279间控制叶片棕色斑点指数的QTL与水稻功能图谱上控制叶绿素含量的QTL的位置一致;表明在亚铁毒胁迫条件下,水稻在其叶片表面出现棕色斑点,叶片衰老,产生一些叶绿素降解物或衍生物,以提高叶片细胞对亚铁等重金属毒害的耐受力。另外,在第11染色体G1465～C950之间检测到了控制叶片棕色斑点指数、茎干重和根干重QTL1个,为主效QTL。在第6染色体XNpb386～XNpb342之间检测到控制茎干重、株高、根长和根干重QTL1个,是否与水稻抗亚铁毒有关需要进一步研究。本研究旨在通过定位与抗亚铁毒有关的QTL,借助与之紧密连锁的分子标记有效地聚合这些QTL,培育出抗亚铁毒性强的水稻新种质材料。     

Multi-Environmental Genetic Analysis of Grain Size Traits Based on Chromosome Segment Substitution Line in Rice (<i>Oryza sativa</i> L.)

Grain size traits are critical agronomic traits which directly determine grain yield, but the genetic bases of these traits are still not well understood. In this study, a total of 154 chromosome segment substitution lines (CSSLs) population derived from a cross between a japonica variety Koshihikari and an indica variety Nona Bokra was used to investigate grain length (GL), grain width (GW), length-width ratio (LWR), grain perimeter (GP), grain area (GA), and thousand grain weight (TGW) under four environments. QTL mapping analysis of six grain size traits was performed by QTL IciMapping 4.2 with an inclusive composite interval mapping (ICIM) model. A total of 64 QTLs were identified for these traits, which mapped to chromosomes 1, 2, 3, 4, 6, 7, 8, 10, 11, and 12 and accounted for 1.6%–27.1% of the total phenotypic variations. Among these QTLs, thirty-six loci were novel and seven QTLs were identified under four environments. One locus containing the known grain size gene, qGL3/GL3.1/OsPPKL1, also have been found. Moreover, five pairs of digenic epistatic interactions were identified except for GL and GP. These findings will facilitate fine mapping of the candidate gene and QTL pyramiding to genetically improve grain yield in rice.     

Detection of quantitative trait loci controlling pre-harvest sprouting resistance by using backcrossed populations of japonica rice cultivars

Backcrossed inbred lines (BILs) and a set of reciprocal chromosome segment substitution lines (CSSLs) derived from crosses between japonica rice cultivars Nipponbare and Koshihikari were used to detect quantitative trait loci (QTLs) for pre-harvest sprouting resistance. In the BILs, we detected one QTL on chromosome 3 and one QTL on chromosome 12. The QTL on the short arm of chromosome 3 accounted for 45.0% of the phenotypic variance and the Nipponbare allele of the QTL increased germination percentage by 21.3%. In the CSSLs, we detected seven QTLs, which were located on chromosomes 2, 3 (two), 5, 8 and 11 (two). All Nipponbare alleles of the QTLs were associated with an increased rate of germination. The major QTL for pre-harvest sprouting resistance on the short arm of chromosome 3 was localized to a 474-kbp region in the Nipponbare genome by the SSR markers RM14240 and RM14275 by using 11 substitution lines to replace the different short chromosome segments on chromosome 3. This QTL co-localized with the low-temperature germinability gene qLTG3-1. The level of germinability under low temperature strongly correlated with the level of pre-harvest sprouting resistance in the substitution lines. Sequence analyses revealed a novel functional allele of qLTG3-1 in Nipponbare and a loss-of-function allele in Koshihikari. The allelic difference in qLTG3-1 between Nipponbare and Koshihikari is likely to be associated with differences in both pre-harvest sprouting resistance and low-temperature germinability.     

Construction of Chromosome Segment Substitution Lines in Peanut (Arachis hypogaea L.) Using a Wild Synthetic and QTL Mapping for Plant Morphology

Chromosome segment substitution lines (CSSLs) are powerful QTL mapping populations that have been used to elucidate the molecular basis of interesting traits of wild species. Cultivated peanut is an allotetraploid with limited genetic diversity. Capturing the genetic diversity from peanut wild relatives is an important objective in many peanut breeding programs. In this study, we used a marker-assisted backcrossing strategy to produce a population of 122 CSSLs from the cross between the wild synthetic allotetraploid (A. ipaënsis×A. duranensis)^4x and the cultivated Fleur11 variety. The 122 CSSLs offered a broad coverage of the peanut genome, with target wild chromosome segments averaging 39.2 cM in length. As a demonstration of the utility of these lines, four traits were evaluated in a subset of 80 CSSLs. A total of 28 lines showed significant differences from Fleur11. The line×trait significant associations were assigned to 42 QTLs: 14 for plant growth habit, 15 for height of the main stem, 12 for plant spread and one for flower color. Among the 42 QTLs, 37 were assigned to genomic regions and three QTL positions were considered putative. One important finding arising from this QTL analysis is that peanut growth habit is a complex trait that is governed by several QTLs with different effects. The CSSL population developed in this study has proved efficient for deciphering the molecular basis of trait variations and will be useful to the peanut scientific community for future QTL mapping studies.     

利用染色体片段置换系定位水稻落粒性主效QTL

水稻落粒性是与其生产密切相关的重要性状之一。以7个染色体片段置换系为材料, 采用重叠群代换作图法对控制落粒性的2个主效QTL进行定位。结果表明, 104个SSR标记在亲本间具有多态性, 多态率为68.0%; 4个置换系的落粒性与亲本日本晴的落粒性相似, 表现难落粒。3个置换系与亲本93-11的落粒性相似, 表现易落粒; 7个染色体片段置换系在第1和第6染色体上检出7个置换片段, 其长度分别为23.6、16.5、 6.6、 9.9、 10.4、 20.2和7.1 cM; qSH-1-1被定位在第1染色体RM472－RM1387之间, 遗传距离约为6.6 cM。qSH-6-1为新发现的落粒性主效QTL, 被定位在第6染色体RM6782－RM3430之间,遗传距离约为4.2 cM。利用染色体片段置换系能准确地定位水稻落粒性QTL, qSH-1-1与qSH-6-1的鉴定和初步定位为其进一步的精细定位、图位克隆及分子标记辅助选择奠定了基础。     

Response of rice to Al stress and identification of quantitative trait Loci for Al tolerance

Rice (Oryza sativa L.) shows the highest tolerance to Al toxicity among small-grain cereal crops, however, the mechanisms and genetics responsible for its high Al tolerance are not yet well understood. We investigated the response of rice to Al stress using the japonica variety Koshihikari in comparison to the indica variety Kasalath. Koshihikari showed higher tolerance at various Al concentrations than Kasalath. The Al content in root apexes was less in Koshihikari than in Kasalath, suggesting that exclusion mechanisms rather than internal detoxification are acting in Koshihikari. Al-induced secretion of citrate was observed in both Koshihikari and Kasalath, however, it is unlikely to be the mechanism for Al tolerance because there was no significant difference in the amount of citrate secreted between Koshihikari and Kasalath. Quantitative trait loci (QTLs) for Al tolerance were mapped in a population of 183 backcross inbred lines (BILs) derived from a cross of Koshihikari and Kasalath. Three putative QTLs controlling Al tolerance were detected on chromosomes 1, 2 and 6. Kasalath QTL alleles on chromosome 1 and 2 reduced Al tolerance but increased tolerance on chromosome 6. The three QTLs explained about 27% of the phenotypic variation in Al tolerance. The existence of QTLs for Al tolerance was confirmed in substitution lines for corresponding chromosomal segments.     

水稻产量及其构成因子对空气CO2浓度增高响应的QTL分析

自由空气CO2浓度增加设施（Free air carbon dioxide enrichment．FACE）使得实际地模拟未来植物生长所处的CO2浓度增加环境变为可能。FACE下．作物生长和产量发生不同程度的加速和提高,而分析作物产量因子对CO2浓度增加响应的遗传基础将有利于对CO2环境变化做出敏感响应的遗传特性的认识,有利于适合未来空气CO2浓度增加环境的高产品种的培育。以粳稻品种Asominori与籼稻品种IR24的杂交组合所衍生的染色体片段置换系（CSSLs）为材料进行田间试验,分别在FACE（约570umol CO2／mol）和正常大气（约370umol CO2／mol）下对籽粒产量及其构成因子等数量性状位点（QTL）进行了分析。结果表明,在FACE下,Asominori和IR24的有效穗数、穗粒数和单株籽粒产量均显著高于对照下的,并且FACE下,65个置换系的变幅范围均大于对照下的;在第1．2,4,6．7,9和12染色体上检测到LOD值在2．5—5．7范围内的控制上述产量性状的20个QTL．其中有3个可以同时在FACE和正常大气下检测到．其余的则只是在某一种CO2环境下检测到。此外,还检测到2个QTL（qFT12 and qGP4）存在着与环境的加性互作效应。可以推论．空气中CO2浓度的增加诱导了部分对CO2浓度敏感的QTL表达,控制水稻产量性状的QTL与CO2增加的环境发生了互作效应。预计利用分子标记辅助育种途径可以培育出适用于未来CO2浓度增加环境下的高产水稻品种。     

利用染色体片段置换系定位水稻落粒性主效QTL

水稻落粒性是与其生产密切相关的重要性状之一。以7个染色体片段置换系为材料,采用重叠群代换作图法对控制落粒性的2个主效QTL进行定位。结果表明,104个SSR标记在亲本间具有多态性,多态率为68.0%;4个置换系的落粒性与亲本日本晴的落粒性相似,表现难落粒。3个置换系与亲本93-11的落粒性相似,表现易落粒;7个染色体片段置换系在第1和第6染色体上检出7个置换片段,其长度分别为23.6、16.5、6.6、9.9、10.4、20.2和7.1 cM;qSH-1-1被定位在第1染色体RM472-RM1387之间,遗传距离约为6.6 cM。qSH-6-1为新发现的落粒性主效QTL,被定位在第6染色体RM6782-RM3430之间,遗传距离约为4.2 cM。利用染色体片段置换系能准确地定位水稻落粒性QTL,qSH-1-1与qSH-6-1的鉴定和初步定位为其进一步的精细定位、图位克隆及分子标记辅助选择奠定了基础。     

覆盖野生稻基因组的染色体片段替换系的构建及其米质相关数量性状基因座位的鉴定

染色体片段替换系（CSSL）是基因组水平快速初步定位数量性状基因座位（QTL）的良好材料,而水稻的品质性状是多基因控制的数量性状,因此可用替换系鉴定控制水稻品质性状的QTL。本文用分子标记辅助选择技术（MAS）构建了由133个株系组成的以‘特青’（籼稻品种）为轮回亲本,以海南的一种普通野生稻为供体亲本,覆盖绝大部分野生稻基因组的染色体片段替换系。利用这套替换系,初步定位了控制稻米外观和理化品质性状的15个QTL,为今后水稻品质性状QTL的克隆以及稻米品质相关性状的改良提供了依据。     

Detection of a quantitative trait locus controlling carbon isotope discrimination and its contribution to stomatal conductance in japonica rice

Increasing leaf photosynthesis offers a possible way to improve yield potential in rice (Oryza sativa L.). Carbon isotope discrimination (Δ¹³C) has potential as an indirect selection criterion. In this study, we searched for quantitative trait loci (QTLs) controlling Δ¹³C, and assessed their association with leaf photosynthesis. Substitution mapping by using chromosome segment substitution lines (CSSLs), that carry segments from the indica cultivar Kasalath in the genetic background of the japonica cultivar Koshihikari, identified genomic regions affecting Δ¹³C on chromosomes (Chr.) 2, 3, 6, 7, and 12. One of the CSSLs, SL208, in which most regions on Chr. 3 were substituted with Kasalath segments, showed higher leaf stomatal conductance for CO₂ (g _s) and Δ¹³C than Koshihikari during the vegetative stage although leaf photosynthetic rate did not differ between them. These results suggest an association between Δ¹³C and g _s. To test this association, we performed a QTL analysis for Δ¹³C at vegetative and heading stages in an F₂ population derived from a cross between SL208 and Koshihikari. The results confirmed a QTL controlling Δ¹³C on the long arm of Chr. 3. By using a near-isogenic line specific to Hd6, we ruled out the possibility that variation in Δ¹³C was generated through the pleiotropic effect of heading date.     

QTL analysis for rice grain length and fine mapping of an identified QTL with stable and major effects

Grain length in rice plays an important role in determining rice appearance, milling, cooking and eating quality. In this study, the genetic basis of grain length was dissected into six main-effect quantitative trait loci (QTLs) and twelve pairs of epistatic QTLs. The stability of these QTLs was evaluated in four environments using an F₇ recombinant inbred line (RIL) population derived from the cross between a Japonica variety, Asominori, and an Indica variety, IR24. Moreover, chromosome segment substitution lines (CSSLs) harboring each of the six main-effect QTLs were used to evaluate gene action of QTLs across eight environments. A major QTL denoted as qGL-3a, was found to express stably not only in the isogenic background of Asominori but also in the recombinant background of Asominori and IR24 under multiple environments. The IR24 allele at qGL-3a has a positive effect on grain length. Based on the test of advanced backcross progenies, qGL-3a was dissected as a single Mendelian factor, i.e., long rice grain was controlled by a recessive gene gl-3. High-resolution genetic and physical maps were further constructed for fine mapping gl-3 by using 11 simple sequence repeat (SSR) markers designed using sequence information from seven BAC/PAC clones and a BC₄F₂ population consisting of 2,068 individuals. Consequently, the gl-3 gene was narrowed down to a candidate genomic region of 87.5 kb long defined by SSR markers RMw357 and RMw353 on chromosome 3, which provides a basis for map-based cloning of this gene and for marker-aided QTL pyramiding in rice quality breeding.