Development and characterization of chromosome segment substitution lines derived from backcross between japonica donor rice cultivar Yukihikari and japonica recipient cultivar Kirara397

Grain yield-related traits and grain quality-related traits are important for rice cultivars. The quantitative trait loci (QTLs) involved in controlling the natural variation in these traits among closely related cultivars are still unclear. The present study describes the development of a novel chromosome segment substitution line (CSSL) population derived from a cross between the temperate japonica cultivars Yukihikari and Kirara397, which are grown in Hokkaido, the northernmost limit for rice cultivation. Days to heading, culm length, panicle length, panicle number, brown grain weight per plant, thousand brown grain weight, brown grain length, brown grain width, brown grain thickness, apparent amylose content, and protein content were evaluated. Panicle length, brown grain length and amylose content differed significantly in the parental cultivars. Thirty-five significant changes in the evaluated traits were identified in the CSSLs. A total of 28 QTLs were located on chromosomes 1, 2, 3, 4, 5, 6, 8, 9, 10, 11 and 12. These findings could be useful for breeding rice cultivars in the northernmost limit for rice cultivation.     

Identification and analysis of QTLs for grain quality traits in rice using an introgression lines population

The demand for superior grain quality represents a major issue in rice (Oryza sativa L.) breeding. To study the genetic basis of rice grain quality, including traits specifying grain dimension and percentage of grains with chalkiness, quantitative genetic analysis was conducted in a population of 37 introgression lines (ILs) of indica elite variety Habataki in the background of japonica cultivar Sasanishiki across two different environments. The ILs showed transgressive segregation for all of the investigated traits, including percentage of grains with chalkiness, brown grain length, width, thickness, length/width ratio, length/thickness ratio, volume and 1,000-brown grain weight, the result indicated that these rice grain quality traits were polygenic phenomenon. Fifty-four QTLs were identified on eleven chromosomes, and 44 QTLs were associated with more than one trait, as a cause of the observed phenotypic correlations between different traits. Five stable QTLs, namely QPGWC.NH-1.1, QPGWC.NH-1.2, QGW.NH-3.1, QLW.NH-7.1 and QGV.NH-8.2, were identified in this study. This study provides useful information for marker-aided improvement of grain quality (grain dimension and percentage of grains with chalkiness) in rice; moreover, ILs carrying these QTLs constitute good candidates for future fine mapping and positional cloning projects.     

Inheritance of the cracking characteristics of abscission layer and its relation with the degree of grain shedding

Seed shattering is an undesirable characteristic of rice cultivars affecting field loss of grain yield before and during harvesting. This research was designed to clarify the inheritance of the cracking characteristics of abscission layers formed between the rice grains and its pedicel using F₂ populations derived from the crossing of Milyang 23 and Ashahi. The results indicated that the cracking characteristic of Milyang 23 was dominant to absence of cracking in Ashahi, and controlled by a single dominant gene. The degrees of grain shedding were somewhat related with the cracking characteristics of the abscission layer. The breaking tensile strengths closely correlated with the diameter of the supporting zone, the thickness of sclerenchyma tissue between the inner side of abscission layer and the central vascular tissues in the lemma side, and in the palea side, respectively. Some negative correlations were observed between the breaking tensile strengths and some agronomic traits such as plant height, culm length, panicle length, number of matured grains, and the ratio of fertility.     

Genotyping by sequencing of rice interspecific backcross inbred lines identifies QTLs for grain weight and grain length

Grain weight and grain length are the most stable components of rice yield and important indicators of consumer preference. Considering the potentials of wild rice and to enhance the rice yields to meet the increasing demands, 185 Backcross Inbred Lines (BILs) in the background of O. sativa ssp. indica cv. PR114, including 63 rufi-BILs derived from O. rufipogon IRGC104433 and 122 glumae-BILs from O. glumaepatula IRGC104387 were evaluated for mapping QTLs for yield and yield component traits using Genotyping by Sequencing (GBS). Phenotypic evaluation of BILs in three seasons spanning two locations revealed significant differences compared with recurrent parent. BILs which did not show significant differences for any trait under investigation, or similar based on pedigree, were excluded from GBS. Some glumae-BILs had to be excluded from mapping QTLs due to less sequence information. A custom designed approach for GBS data analysis identified 3322 informative SNPs in 55 rufi-BILs and 3437 informative SNPs in 79 glumae-BILs. QTL mapping identified one QTL for thousand grain weight (qtgw5.1), two for grain width (qgw5.1, qgw5.2) and one for grain length (qgl7.1) in rufi-BILs. In the glumae-BILs, three QTL for thousand grain weight (qtgw2.1, qtgw3.1, qtgw6.1) and two for grain length (qgl3.1, qgl7.1) were identified. Most of the grain weight and width QTL showed positive additive effect contributed by wild species allele, whereas the grain length QTL showed positive additive effect contributed by recurrent parent allele. Based on their physical position, none of the QTLs were found similar to previously cloned QTLs. QTLs for grain traits identified from low yielding wild relatives of rice reveals their significance in improving further the rice yields and widen the genetic base of cultivated rice.     

Haplotype structure in grain weight gene GW2 and its association with grain characteristics in rice

GW2, a grain weight quantitative trait locus (QTL) in rice encodes a ring type E-3 ubiquitin ligase. A single nucleotide deletion at the 346th nucleotide position in the ligase domain of GW2 was earlier reported to result in higher grain weight in rice. The present study aimed at validating the known functional polymorphism and identifying additional natural genetic variation if any, in the region that included the functional domain of GW2 in a set of indica and aromatic genotypes for which ninety three rice genotypes were phenotyped for grain length, grain width and 100 grain weight. A wide range of variation was observed for these traits. PCR amplification and sequencing of GW2 target region revealed absence of insertion/deletion (InDel) at the 346th position which suggested that the genetic variation in grain weight in Basmati and non-Basmati indica genotypes was not explained by this InDel. However, four new single nucleotide polymorphisms (SNPs) were discovered at nucleotide positions 406, 461, 466 and 501 in the fifth exon and one InDel each in second and fourth introns. Only two of these SNPs, at positions 461 and 501 led to amino acid substitutions. A total of 10 haplotypes were constructed based on these four SNPs which could be regrouped into four categories based on their amino acid substitutions. Association genetic analysis of these haplotypes with different grain traits revealed a moderate association with grain width (R² = 0.18 at P < 0.05). Thirteen haplotypes constructed using both intronic and exonic polymorphisms did not have any association with grain traits.     

水稻转录因子bHLH家族基因响应环境胁迫表达谱分析

植物转录因子在植物的生长发育及其对外界环境的反应中起着重要的作用。bHLH转录因子是植物转录因子最大的家族之一。本研究以水稻日本晴为材料,在苗期进行PEG模拟干旱胁迫和ABA（abscisicacid）处理,采用荧光定量PCR方法,比较分析干旱胁迫差异表达的22个候选bHLH家族基因在PEG和ABA处理下的不同时间梯度的表达谱。结果表明大部分候选基因的PEG胁迫表达谱与干旱胁迫表达谱一致,验证了芯片结果的可靠性;而在ABA处理下的大部分bHLH基因表现为抑制表达,部分基因的表达谱与干旱或PEG胁迫表达谱明显不同,推测这些bHLH基因参与PEG和ABA胁迫应答的具有相同或不同的分子路径或模式。研究结果对进一步克隆这些基因及分析其在植物生长发育过程中的分子调控模式打下了基础。     

Identification and characteristics of quantitative trait locus for grain protein content, <Emphasis Type="Italic">TGP12</Emphasis>, in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Grain protein content is an important analysis target to determine grain quality in rice. This study analyzed quantitative trait loci (QTLs) for the content of grain protein and amylose using the chromosomal segment substitution lines developed from ‘Koshihikari’ and ‘Nona Bokra’. It also evaluated the effects of target QTL on eating and cooking quality through the physical properties of cooked rice and its gel consistency. QTL analysis over 3 years detected the QTL on chromosome 12, TGP12, which consistently decreased total grain protein content via the ‘Nona Bokra’ allele. Selected CSSL with TGP12, CSSL-TGP12, showed a lower content of total grain protein in brown and milled rice, and had similar amylose content, grain size, and weight of brown rice, compared with ‘Koshihikari’. Based on the results of sodium dodecyl sulfate polyacrylamide gel electrophoresis, brown rice with CSSL-TGP12 had no remarkable decrease or loss in any specific protein. Regarding eating and cooking quality, CSSL-TGP12 did not show stable effects on physical properties, hardness, stickiness, or adherability of cooked rice or its gel consistency. These results suggest that TGP12 could be one of the key genetic factors for the alteration of grain protein content without an effect on eating or cooking quality.     

Development of photoperiod- and thermo-sensitive male sterility rice expressing transgene Bacillus thuringiensis

Stem borers and leaffolders are the main pests that cause severe damage in rice (Oryza sativa L.) production worldwide. We developed the first photoperiod- and thermo-sensitive male sterility (PTSMS) rice 208S with the cry1Ab/1Ac Bacillus thuringiensis (Bt) gene, through sexual crossing with Huahui 1 (elite line with the cry1Ab/1Ac gene). The novel 208S and its hybrids presented high and stable resistance to stem borers and leaffolders, and the content of Cry1Ab/1Ac protein in chlorophyllous tissues achieved the identical level as donor and showed little accumulation in non-chlorophyllous tissue. No dominant dosage effect in the Bt gene was observed in 208S and its derived hybrids. An analysis of fertility transition traits indicated that 208S was completely sterile under long day length/high temperature, but partially fertile under short day length/low temperature. With fine grain quality and favorable combining ability, 208S had no observed negative effects on fertility and agronomic traits from Bt (cry1Ab/1Ac). Additionally, 208S as a male sterile line showed no fertility decrease caused by Bt transgenic process, as it is the case in Huahui 1. Thus, 208S has great application value in two-line hybrid production for insect resistance, and can also be used as a bridge material in rice Bt transgenic breeding.     

水稻早恢402体细胞无性系及其杂种F1的粒形与碾米品质

比较了水稻早熟恢复系402的体细胞无性系及其杂种F1的粒形及碾米品质的变异特点。结果表明，R3株系与供体亲本籽粒间及体细胞组培杂种R1F2与亲本杂种F2籽粒间米质性状的总体差异达极显著水平。R3的粒宽、粒厚、糙米率、精米率倾小值变异；R1F2的粒长、长/宽比、长/厚比及整精米率等指标倾大值变异。R3代的糙米率及精米率与整     

不同氮肥水平下中熟籼稻和粳稻产量、氮素吸收利用差异及相互关系

选用生育期相近的中熟中籼(138~143 d)和中熟中粳(136~145 d),于大田条件下探讨了4种氮肥水平即0、150、225、300 kg hm^-2纯氮对产量的影响及不同生育阶段氮素吸收利用的差异。结果表明,随着氮肥施用量的增加粳稻平均产量呈增加趋势,而籼稻平均产量中氮水平达最高,高氮水平有所降低。各处理下籼稻产量明显高于粳稻,籼稻有较高的穗粒数和千粒重,平均分别比粳稻高22.45%、7.51%。植株氮素阶段吸收量,拔节、抽穗和成熟期籼稻平均分别比粳稻高24.26%、3.14%和6.45%;植株氮素阶段吸收速率,移栽至拔节、拔节至抽穗和抽穗至成熟阶段籼稻平均分别比粳稻高38.07%、16.05%和23.22%。相关分析表明,籽粒产量与氮素阶段吸收量、阶段吸收速率和氮肥利用率呈显著或极显著正相关关系。说明生育期相近的中熟中籼产量高于中熟中粳与籼稻植株较强的氮素吸收特性有关。     

Genetic dissection of rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) tiller,plant height,and grain yield based on QTL mapping and metaanalysis

Tiller number per plant (TN) and plant height (PH) are important agronomic traits related to grain yield (GY) in rice (Oryza sativa L.). A total of 30 additive quantitative trait loci (A-QTL) and 9 significant additive × environment interaction QTLs (AE-QTL) were detected, while the phenotypic and QTL correlations confirmed the intrinsic relationship of the three traits. These QTLs were integrated with 986 QTLs from previous studies by metaanalysis. Consensus maps contained 7156 markers for a total map length of 1112.71 cM, onto which 863 QTLs were projected; 78 meta-QTLs (MQTLs) covering 11 of the 30 QTLs were detected from the cross between Dongnong422 and Kongyu131 in this study. A total of 705 predicted genes were distributed over the 21 MQTL intervals with physical length <0.3 Mb; 13 of the 21 MQTLs, and 34 candidate genes related to grain yield and plant development, were screened. Five major QTLs, viz. qGY6-2, qPH7-2, qPH6-3, qTN6-1, and qTN7-1, were not detected in the MQTL intervals and could be used as newly discovered QTLs. Candidate genes within these QTL intervals will play a meaningful role in molecular marker-assisted selection and map-based cloning of rice TN, PH, and GY.     

Pleiotropic effects of the elongated glume gene P1 on grain and spikelet shape-related traits in tetraploid wheat

Wheat grain size and shape are associated not only with yield but also with product and milling quality. A subspecies of cultivated tetraploid wheat, Triticum turgidum ssp. polonicum, is characterized by elongated glumes. To elucidate morphological effects of the subspecies differentiation-related gene, we conducted QTL analysis for grain and spikelet shape using a mapping population between two tetraploid wheat subspecies, polonicum and durum. P1, the gene controlling the elongated glumes, was located on chromosome 7A, and the polonicum-type allele acted in an incomplete dominance manner to express the elongated glume phenotype. The polonicum allele of the P1 locus significantly affected not only glume length but also grain shape, spike shape, awn length and seed fertility in tetraploid wheat. The elongated glume phenotype was correlated with an increase in spike length, grain length and grain weight, and with a decrease in fertility, grain number and awn length. Thus, the subspecies differentiation-related gene in subspecies polonicum dramatically affects grain shape accompanied by alteration of spikelet shape in tetraploid wheat.     

Rice tungro spherical virus resistance into photoperiod-insensitive japonica rice by marker-assisted selection

Rice tungro disease (RTD) is one of the destructive and prevalent diseases in the tropical region. RTD is caused by Rice tungro spherical virus (RTSV) and Rice tungro bacilliform virus. Cultivation of japonica rice (Oryza sativa L. ssp japonica) in tropical Asia has often been restricted because most japonica cultivars are sensitive to short photoperiod, which is characteristic of tropical conditions. Japonica1, a rice variety bred for tropical conditions, is photoperiod-insensitive, has a high yield potential, but is susceptible to RTD and has poor grain quality. To transfer RTD resistance into Japonica1, we made two backcrosses (BC) and 8 three-way crosses (3-WC) among Japonica1 and RTSV-resistant cultivars. Among 8,876 BC₁F₂ and 3-WCF₂ plants, 342 were selected for photoperiod-insensitivity and good grain quality. Photoperiod-insensitive progenies were evaluated for RTSV resistance by a bioassay and marker-assisted selection (MAS), and 22 BC₁F₇ and 3-WCF₇ lines were selected based on the results of an observational yield trial. The results demonstrated that conventional selection for photoperiod-insensitivity and MAS for RTSV resistance can greatly facilitate the development of japonica rice that is suitable for cultivation in tropical Asia.     

Effect of panicle nitrogen on grain filling characteristics of high-yielding rice cultivars

The effects of four levels of panicle nitrogen fertilizer on the grain filling characteristics of three rice (Oryza sativa L.) varieties (super japonica rice Ningjing 3, three-line japonica hybrid rice Changyou 3 and three-line indica–japonica hybrid rice Yongyou 12) were analyzed. The results showed that difference in time of maximum filling rate (T_max) was the smallest between the superior and inferior spikelets of Ningjing 3, bigger between those of Changyou 3, the biggest between those of Yongyou 12. Ningjing 3 was of the synchronous grain-filling type, Yongyou 12 the asynchronous type, and the Changyou 3 the medium type. The grain filling rate, the initial filling power (R₀), the maximum filling rate (G_max), and the average filling rate (G) of the superior and inferior spikelets of the three varieties under the treatment of panicle N with the amount of 120 kg ha⁻¹ (middle panicle N, shortened as NM) treatment were higher than those of other treatments. NM treatment led to the highest increase in grain weight at the middle stage of filling for all the three varieties. The treatment shortened the early and late stages of grain filling, but extended the middle stage when the filling rate was the highest. However, the middle stage of grain filling of Changyou 3 and Yongyou 12 was much more extended than that of Ningjing 3, indicating a better effect of N on hybrid rice varieties.     

Quantitative trait loci for grain-quality traits across a rice F2 population and backcross inbred lines

Improving grain-quality is an important goal in rice breeding programs. One vital step is to find major quantitative trait loci (QTLs) for quality related traits and then investigate the relationships among them. We crossed ‘N22’, an indica variety with good appearance but low grain weight, to a japonica variety, ‘Nanjing35’, with superior grain yield but poor appearance. This enabled us to construct an F₂ population and a set of backcross inbred lines (BILs) for QTL-mapping for the traits related grain appearance. In all, 37 QTLs were identified for grain length (GL), grain width (GW), grain thickness (GT), thousand-grain weight (TGW), and the percentage of grains with chalkiness (PGWC). Of these, 17 QTLs detected from 184 plants in the F₂ population explained 4.97–27.26 % of the phenotypic variance, another 20 QTLs were identified using BILs from 2009 to 2010. Quantitative trait loci for major effects were detected in different populations and across years. A new QTL hot spot (marker interval RM504–RM520) was found on Chromosome 3, which harbored QTLs for GL, GW, GT, and TGW. Among our five examined traits, grain shape was significantly correlated with TGW and PGWC. The PGWC values of two heavier grains BILs, L93, and L145 are much lower than Nanjiing35, the analysis of genotype showed that this greater weight may due to the locus for GL occurring within RM504–RM520 on Chromosome 3. Therefore, those two lines will allow us to develop a long-grain high-yield rice variety with less chalkiness.     

Identification and application of major quantitative trait loci for panicle length in rice (Oryza sativa) through single‐segment substitution lines

Panicle length (PL), an important yield‐related trait, strongly affects yield components, such as grain number, grain density and rice quality. More than 200 panicle length quantitative trait loci (PL QTLs) are identified, but only a small number are applied in rice breeding. In this study, we performed QTL analysis for PL using 42 single‐segment substitution lines (SSSLs) derived from nine donors in the genetic background of HJX74. Fourteen QTLs and five heterosis QTLs (HQTLs) for PL were recognised. Three QTLs and four HQTLs acted positively, and the other eleven QTLs and one HQTL acted negatively. By scanning the single heterozygous background region of the F₂ population with large‐genetic‐effect SSSLs, we mapped PL loci qPL6‐2 and qPL7‐1 to different locations on chromosomes 6 and 7, respectively, in three consecutive years of independent trials. The genetic effects of these QTLs were further assessed. qPL6‐2 demonstrated the most positive additive effect (QTL), whereas qPL7‐1 achieved the most positive dominant effect (HQTL) for PL. These results indicated that the pyramiding of PL QTLs might increase grain yield and facilitate the application of the beneficial allele in hybrid rice breeding.     

Genome‐wide association analysis reveals flowering‐related genes regulating rachis length in rice

Rachis length is correlated with panicle size in rice. Unveiling the genetic basis of rachis length is important for understanding the genetic regulation of panicle size. In this study, we performed a genomewide association study of rachis length using 529 rice accessions from two environments. In total, 20 loci were identified for rachis length and distributed across the 12 chromosomes except for chromosomes 2 and 5. Thirteen of the 20 loci were not linked to the cloned panicle size genes. Six flowering‐related genes, including the CCT domain‐containing genes Ghd7, Ghd7.1, Hd1, OsCCT1 and OsCO3 as well as Ehd1, were associated with rachis length in this study. These findings suggest that the network of these flowering‐related genes probably participates in the regulation of rachis length and thus affects panicle length and yield. Interestingly, haplotype analysis showed that OsCCT1 is a putative candidate gene which plays a key role in the variation of rachis length. Together, the genetic loci identified in this study could be used for rice breeding by marker‐assisted selection to improve grain yield.     

利用BC2F2高代回交群体定位水稻籽粒大小和形状QTL

以我国优良籼稻恢复系蜀恢527为轮回亲本, 以来自菲律宾的Milagrosa为供体亲本, 培育了样本容量为199株的BC₂F₂高代回交群体。选取85个均匀分布在12条染色体上的多态性SSR标记进行基因型分析, 同时对粒长、粒宽、长宽比和千粒重4种性状进行了表型鉴定。采用性状－标记间的单向和双向方差分析对上述性状进行了QTL定位。单向方差分析(P<0.01)共检测到了10个控制粒长、粒宽、长宽比和千粒重的QTL, 其中有3个具有多效性。由于粒长和长宽比的高度相关性, 控制长宽比的2个QTL均能在粒长QTL中检测到。位于第3染色体着丝粒区域的qgl3b是一个控制粒长、长宽比和千粒重的主效QTL, 它可以分别解释粒长、长宽比和千粒重表型变异的29.37%、26.15%和17.15%。该QTL对于粒长、长宽比和千粒重均表现较大的加性效应(来自蜀恢527的等位基因为增效)和负向超显性。位于第8染色体的qgw8位点是一个控制粒宽的主效QTL, 同时也是控制千粒重的微效QTL, 能解释粒宽表型变异的21.47%和千粒重表型变异的5.16%。该QTL对粒宽和千粒重均具有较大的加性效应(来自蜀恢527的等位基因为增效)和正向部分显性。双向方差分析(P<0.005)共检测到61对显著的上位性互作, 涉及54个QTL, 其中23个是能同时影响2~4个性状的多效位点, 且有8个位点与单向方差分析检测到的相同。控制长宽比的13对上位性互作位点中, 与控制粒长的上位性互作位点完全相同的有8对。以上结果为进一步开展水稻籽粒大小和形状有利基因的精细定位、克隆和分子设计育种奠定了基础。     

Advanced backcross QTL analysis reveals complicated genetic control of rice grain shape in a japonica × indica cross

Grain shape is an important trait for improving rice yield. A number of quantitative trait loci (QTLs) for this trait have been identified by using primary F₂ mapping populations and recombinant inbred lines, in which QTLs with a small effect are harder to detect than they would be in advanced generations. In this study, we developed two advanced mapping populations (chromosome segment substitution lines [CSSLs] and BC₄F₂ lines consisting of more than 2000 individuals) in the genetic backgrounds of two improved cultivars: a japonica cultivar (Koshihikari) with short, round grains, and an indica cultivar (IR64) with long, slender grains. We compared the ability of these materials to reveal QTLs for grain shape with that of an F₂ population. Only 8 QTLs for grain length or grain width were detected in the F₂ population, versus 47 in the CSSL population and 65 in the BC₄F₂ population. These results strongly suggest that advanced mapping populations can reveal QTLs for agronomic traits under complicated genetic control, and that DNA markers linked with the QTLs are useful for choosing superior allelic combinations to enhance grain shape in the Koshihikari and IR64 genetic backgrounds.     

Relationship between grain yield and quality in rice germplasms grown across different growing areas

Rice grain yield and quality are two major foci of rice breeding. In this study, Chinese regional rice test data provide us the unique opportunity to analyze the relationship between yield and quality in rice, because China has an unusually wide range of rice cultivars. We analyzed the relationships between grain yield, yield components, and grain quality of 300 rice germplasms. Japonica was superior in both yield and quality compared with indica. A high setting rate improved the head rice ratio. A higher 1000 grain weight was negatively correlated with quality characteristics but had a positive correlation with yield. A high spikelet density (number of grains per centimeter on the panicle) not only benefits the yield but also the head rice ratio and chalkiness traits. According to our results, global rice production can be increased to at least 8500 kg/ha to meet projected demands in 2025 without sacrificing grain quality.