Chromosomal location of fertility restoring genes for ‘wild abortive’ cytoplasmic male sterility using primary trisomics in rice

Summary Identification and location of fertility restoring genes facilitates their deployment in a hybrid breeding program involving cytoplasmic male sterility (CMS) system. The study aimed to locate fertility restorer genes of CMSWA system on specific chromosomes of rice using primary trisomics of IR36 (restorer), CMS (IR58025A) and maintainer (IR58025B) lines. Primary trisomic series (Triplo 1 to 12) was crossed as maternal parent with the maintainer line IR58025B. The selected trisomic and disomic F₁ plants were testcrossed as male parents with the CMS line IR58025A. Plants in testcross families derived from disomic F₁ plants (Group I crosses) were all diploid; however, in the testcross families derived from trisomic F₁ plants (Group II crosses), some trisomic plants were observed. Diploid plants in all testcross families were analyzed for pollen fertility using 1% IKI stain. All testeross families from Group I crosses segregated in the ratio of 2 fertile: 1 partially fertile+partially sterile: 1 sterile plants indicating that fertility restoration was controlled by two independent dominant genes: one of the genes was stronger than the other. Testcross families from Group II crosses segregated in 2 fertile: 1 partially fertile+ partially sterile: 1 sterile plants in crosses involving Triplo 1, 4, 5, 6, 8, 9, 11 and 12, but families involving triplo 7 and triplo 10 showed significantly higher X² values, indicating that the two fertility restorer genes were located on chromosome 7 and 10. Stronger restorer gene (Rf-WA-1) was located on chromosome 7 and weaker restorer gene (Rf-WA-2) was located on chromosome 10. These findings should facilitate tagging of these genes with molecular markers with the ultimate aim to practice marker-aided selection for fertility restoration ability.     

High-lysine mutants of rice, Oryza sativa L.

In order to improve the nutritive value of rice protein, it is necessary to increase the level of lysine and threonine. Ten high-lysine mutants were selected from N-methyl-N-nitrosourea-induced (MNU-induced) mutant lines, using high-performance liquid chromatography for amino acid analysis. The lysine content of these mutants ranged from 5.10% to 6.38% of total amino acids, while that of the original varieties was about 4%. All high-lysine mutants also had a greatly increased histidine content and a floury endosperm. An increase in lysine content of the albumin/globulin fraction was characteristic of the MNU-induced high-lysine mutants. The segregation ratio of normal to floury endosperm in the F2 seeds from a cross between ‘Kinmaze’ and a high-lysine mutant fitted the expected 3:1 ratio. The high level of lysine and the floury endosperm may be controlled by the same gene.     

Chromosomal regions controlling anther culturability in rice (Oryza sativa L.)

Diallel analysis has revealed that anther culturability in rice (Oryza sativa L.) is a quantitative trait controlled by the nuclear genome. Mapping of anther culturability is important to increase the efficiency for green plant regeneration from microspores. In the previous study, we detected distorted segregation of RFLP markers in rice populations derived from the anther culture of an F₁ hybrid between a japonica cultivar ‘Nipponbare’ and an indica cultivar ‘Milyang 23’. To clarify the association between chromosomal regions showing distorted segregation and anther culturability, the anther culturability of doubled haploid lines derived from the same cross combination was examined, and the association between alleles of the RFLP markers, which exhibiting the most distorted segregation on chromosomes 1, 3, 7, 10 and 11, and the anther culturability was evaluated. One region on chromosome 1 was found to control callus formation from microspores, and one region on chromosome 10 appeared to control the ratio of green to albino regenerated plants. In both regions, the Nipponbare allele had positive effects. Three regions on chromosomes 3, 7 and 11, however, showed no significant effect on anther culturability. This revised version was published online in August 2006 with corrections to the Cover Date.     

水稻高秆突变体株高及其构成的比较和等位性分析

利用伽玛射线辐射诱变获得了2份高秆突变体Gh-2和Sh-3,对它们的株高及其各节间长、穗长等的变化进行了分析.以同样是高秆的突变体Mh-1做测验种与Gh-2和Sh-3进行杂交,研究了相互间的等位性,研究结果认为它们是互为等位的遗传关系,即含有对sd-1能起隐性调节作用的i-sd1(t)基因.     

Genetic diversity among parental lines of Indica hybrid rice (Oryza sativa L.) in China based on coefficient of parentage

Genetic diversity constitutes the raw material for plant improvement, and provides protection against genetic vulnerability to biotic and abiotic stresses. Diversity of parental lines of indica hybrid rice in China is not well‐characterized. The major objective of this study was to quantify genetic diversity of Chinese parental lines of hybrid rice via coefficient of parentage (COP). All 100 parental lines of hybrid rice widely used in hybrid breeding and commercial production during 1976–2003 were studied by COP analysis. The mean COP for the 100 parental lines was low (0.056), indicating a potentially high degree of diversity in Chinese hybrid rice breeding. Forty‐nine percent of all pairs of parental lines were completely unrelated by pedigree data. The low mean COP for the parental lines was attributed to a continual incorporation of exotic germplasm (wild rice, japonica and javanica etc.) into the genetic base over time, to the introduction of foreign germplasm from the Philippines (International Rice Research Institute), Korea, the United States, Thailand, and Guyana as breeding stock. The mean COP from 1976 to 1990 was twice as much as that from 1990 to 2003. Cluster analysis was an effective method to discriminate diversity, ten clusters were identified, and maintainer lines, restorer lines and other parental lines with special genetic background were clearly grouped. In addition, restorer lines were further divided into 11 sub‐clusters, which basically was in agreement with hybrid rice breeding. Among ten provinces, Hunan, Sichuan and Fujian were outstanding for breeding 54 of 100 parental lines in hybrid rice production, and the genetic diversity of parental lines in Fujian, Sichuan,Guangxi, Hunan and Jiangsu were all narrower than that in Hubei, Guangdong, Zhejiang and Jiangxi. The result of coefficient of parentage analysis for 100 parental lines may promote the management of parental diversity and hybrid rice breeding in China.     

Complex constitutive nature of Japanese upland rice, Oryza sativa L.

Two rice ecotypes, the so-called lowland and upland populations, which carry different isozyme genotypes mostly at a single locus, are cultivated in Japan. The aim of this study was to examine the origin and the mechanism for keeping these genetic differences. The upland population is cultivated in upland fields and carries a different allele for a particular isozyme gene, Pgd-1, which has never been found in the lowland population. RFLP markers showed a weak trend for genetic differentiation between the two ecotypes. On the other hand, morphological, and physiological traits showed marked differences between the two ecotypes. Furthermore, based on the genotypic difference, two Japonica subgroups are defined in the upland population. Subgroup I is the minor group and carries key lowland characters, including the genotype for PGD. Subgroup II carries different traits and the genotype for PGD of the alternative subgroup. As an allelic difference for Pgd-1 is known to occur between the two ecospecies, Tropical (Tr) and Temperate (Tm) Japonicas, upland cultivars can be classified by diagnostic characters which distinguish a variety into Tr or Tm type. The upland population consists of three types of cultivars, Tr-, Tm- and intermediate-type. In contrast, the lowland population consists of a uniform Tm type Japonicas. As Japanese upland cultivars still have an isozyme allele specific to the Tr type, the upland population has a rather complex constitution which is presumably now being introgressed by lowland genetic material, but still represents a major difference at some genetic levels. Upland rice carries several stress-resistant genes which would be useful for lowland rice breeding. The genetic difference would be efficient for tagging upland specific traits by upland specific genetic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytogenetical studies on African rice,Oryza glaberrima Steud. 3. Primary trisomics produced by pollinating autotriploid with diploid

Summary In order to promote cytogenetical studies on Oryza glaberrima (2n=2x=24), I produced primary trisomics in the progeny of the autotriploid. When the autotriploid was pollinated with the diploid, 136F1 seeds (5.8% of the number of spikelets pollinated) were obtained. The numbers of chromosomes of 31 surviving progeny varied from 24 to 28, with plants having 2n=25 occurring at the highest frequency (34.4%). The primary trisomics differed from one another as well as from the normal disomics in many aspects. The differences were attributed to a single extra chromosome in each trisomic type. Nine primary trisomics were classified into 8 types on the basis of morphological and reproductive features. Seeds were obtained from all primary trisomics except for Type D by self pollination, open pollination or pollination with the disomics. I was able to maintain 7 types of primary trisomics by seed propagation. The primary trisomics which are capable of expressing subtle genetic dissimilarities should be useful for studying the relationship between the two cultivated species of rice at the level of individual chromosomes.     

Molecular mapping of a new genic male-sterility gene causing chalky endosperm in rice ( Oryza sativa L.)

A genic male-sterility gene newly induced by chemical mutagenesis, tentatively designated as ms-h(t), was located on the molecular map of rice and tested for its effect on chalky endosperm. Bulked segregant analysis was used to determine the chromosomal location of the ms-h(t) locus by screening four to five RFLP markers per chromosome. After confirming that the gene was located on chromosome 9, twenty-four RFLP markers from chromosome 9 were surveyed for polymorphism in the parents of the mapping population. Of these, eleven markers were mapped around the ms- h(t) locus. RG451 and RZ404 flanked the ms-h(t) gene, at 2.5cM and 3.3cM, respectively. Heterozygous F2 to F4 progenies were tested for co- segregation of male-sterility and chalky endosperm and it was revealed that ms-h(t) might have a pleiotropic effect on chalky endosperm. This mutant would be a good biological material to characterize the biochemical mechanism of male sterility and related pleiotropic effects. Further studies should be needed to know the usefulness of this mutant for hybrid seed production. This revised version was published online in July 2006 with corrections to the Cover Date.     

Trisomic analysis of a strong photoperiod-sensitivity gene E1 in rice (Oryza sativa L.)

Recent genetic analyses on heading-time of rice indicated that almost all the well-adapted varieties in the temperate zone carry a strong photoperiod-sensitivity gene E1, a dominant allele of E1 locus. In order to identify the chromosome on which E1 is located, a trisomic analysis was made using two primary trisomic series originating from the japonica varieties, Nipponbare and Kinmaze, respectively. The Nipponbare and Kinmaze series were crossed with heading-time tester lines, EG0 and EG3, respectively, both of which did not carry the E1. The F2 populations for chromosome 1, 2, and 3 could not be analyzed due to lack of seed. All the other F2 populations showed distinct segregation into early-type and late-type plants caused by the E1 locus segregation, which suggested that the trisomic analysis for E1 locus could be efficiently made. Both disomic and trisomic groups in the F2 population from the cross of the trisomic line for chromosome 7 × EG0 showed a segregation ratio significantly different at the 1% level from a ratio of 1 [e1e1; early]: 3 [E1e1, E1E1; late]. This suggested that E1 was located on chromosome 7. Subsequently, the linkage analysis was made using three morphological marker genes on chromosome 7. It was recognized that E1 was linked to rfs (rolled fine strip gene) and slg (slender glume gene) with recombination values of 16.3 ± 5.88 and 9.1 ± 4.72%, respectively. From these results, it can be concluded that E1 is most likely to be located on chromosome 7. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fine mapping and photosynthetic characteristics of the lower chlorophyll b 1 mutant in rice (Oryza sativa L.)

Chlorophylls absorb and transfer light energy to the photosynthetic system. Consequently, chlorophyll content is strongly related to crop biomass and yield. We isolated a rice spontaneous mutant, lower chlorophyll b 1 (lcb1), from a recombinant inbred line population. Under normal growth conditions, lcb1 plants produced yellow leaves with decreased total chlorophyll and chlorophyll b contents, but normal chlorophyll a content. Photosynthetic and fluorescence parameters differed between wild‐type and lcb1 plants. Compared with wild type, lcb1 had a higher electron transfer rate, a lower photochemical quenching coefficient and significantly reduced grain number, biomass and yield. A recessive nuclear gene controlled the mutant trait. Through map‐based cloning, we located the LCB1 gene in a 117.4‐kb region on the short arm of chromosome 3, close to the centromere, in a region containing 15 predicted candidate genes. None of these genes was directly related to chlorophylls or the chloroplast; therefore, lcb1 may be a mutation of a novel gene. These results will be useful for further research on the molecular mechanisms controlling biogenesis and chloroplast biochemical processes.     

AMMI analysis of rainfed lowland rice (Oryza sativa) trials in Nigeria

The additive main effects and multiplicative interaction (AMMI) model is used to analyse the grain yield data of 13 rice genotypes grown in 12 rainfed lowland rice environments. The trials were organized by the International Network for Genetic Evaluation of Rice in Africa (INGER-Africa) and conducted in Nigeria. Main effects due to environments (E), genotypes (G) and G × E interaction were found to be significant (P = 0.001). Cross validation analysis suggested that an AMMI model with one interaction principal component axis (IPCA) was most useful predictively, whereas Gollobs’ test declared two components, IPCA1 and IPCA2, statistically significant (P = 0.01). The IPCAl, however, accounted for most (47.8%) of the G × E sum of squares. Correlation and regression analysis, and relative scatter of genotype and environment points on the AMMI biplot suggest that the interaction partitioned in IPCA1 resulted from differences in the days to flowering among the genotypes. The paper discusses these in relation to the occurrence of Fe toxicity at the test sites and varietal tolerance to the stress.     

Variation among progenies of diploid plants regenerated from haploid, microspore-derived cell-suspension protoplasts of rice (Oryza sativa L.)

Progenies of 110 diploid R₀ plants regenerated from haploid protoplasts derived from microspore callus cell suspension culture of rice (cv. Miara) were evaluated for 6 traits, along with control lines, in a replicated field trial. Complementary post-harvest observations were carried out for four panicle or grain characters. A wide range of variation was observed for all traits under study among the protoclonal lines. The mean of the protoclonal population was significantly different from that of the control for most traits and 64.5% of the lines differed from the control for at least one field-evaluated trait. A unidirectional shift towards later-maturing, taller, and lower-tillering lines was observed. Of all the lines, 39% were taller than the control by at least 10 cm, 31 % had flowering delayed by at least 5 days, and 13% exhibited significantly longer panicles (P = 0.05). As to the agronomical value of the lines, the variation resulted in a global negative drift although favourable variations were found in a few lines. All but two lines were homogeneous, suggesting that most variations existed in a homozygous form in the R₀ plants. R₂ lines grown from seeds of 17 R₁ protoclonal lines stably inherited the changes and did not segregate, thereby confirming that variations are fixed.     

Crosses between cultivars and tissue culture-selected plants for salt resistance improvement in rice, Oryza sativa

Several R₂ somaclonal families were derived from plants regenerated from a salt‐resistant callus of the salt‐sensitive rice cultivar ‘I Kong Pao’ (IKP). The family R₂‐1‐23, in the presence of NaCl exhibited higher yield performances than the initial cultivar. This improvement in salinity resistance, however, was not transmitted to following generations; despite a higher number of spikelets per plant, family R₃‐1‐23 did not perform better than the initial cultivar because of a very low seed set. This somaclonal family, its initial being the cultivar IKP, the breeding line IR₃1785 (extremely salt‐sensitive) and the cultivar ‘Aiwu’ (moderately salt‐resistant), were used as parents for production ofhybrids. Four crosses, IKP×R3‐1‐23, IR31785 ×R3‐1‐23, IR31785× IKP and IKP בAiwu’, were performed. Most of the F₁ hybrids cultivated in the absence of salt exhibited increased performances compared with the mid‐parent, suggesting an heterosis effect for yield‐related parameters. F₂ populations were screened for salinity resistance and a clear improvement for yield in stress conditions was recorded for populations derived from IK×R3‐1‐23, IR31785×R3‐1‐23 and IR31785×IKP, although the mean level of increase over the mid‐parent (RIMP) varied depending on the population, the presence or absence of stress, and the quantified parameters. The results are discussed in relation to the usefulness of in vitro selection for obtaining interesting somaclonal variants useful to be integrated in classical breeding programmes for salinity resistance in rice.     

水稻温敏型叶片白化转绿突变体tsa2的表型鉴定与基因定位

水稻温敏型叶色突变体是研究植物光合作用、叶绿体结构和功能以及温度影响叶绿体发育的理想材料。利用甲基磺酸乙酯(EMS)诱变籼型水稻(OryzasativaL.)三系保持系西农1B,从其后代中筛选到一个突变性状稳定遗传的温敏型叶片白化转绿突变体tsa2 (temperature-sensitive green-revertible albino 2)。与野生型相比, tsa2突变表型受温度影响, 22°C条件下萌发的野生型幼苗表型正常,而tsa2幼苗完全白化,且约40%白化苗死亡,存活白化苗的光合色素含量、光合速率均显著降低,成熟期主要农艺性状均显著变劣;在28°C下萌发的tsa2幼苗叶片呈浅绿色并伴有白条纹,其光合色素含量显著降低,光合速率及主要农艺性状差异较小; 32°C下萌发的tsa2幼苗叶片无明显差异。透射电镜观察显示,与野生型相比,tsa2在22°C下叶肉细胞中无叶绿体或存在异常发育叶绿体(尚未分化出基粒和基层),在28°C下部分叶肉细胞含少量发育完整的叶绿体,在32°C下叶肉细胞数量及形态均正常。实时荧光定量PCR(qRT-PCR)分析表明,与野生型相比,tsa2突变体中部分光合色素代谢途径基因、叶绿体发育相关基因及光合作用相关基因的表达水平呈不同程度变化。遗传分析表明, tsa2突变表型受一对隐性核基因控制, TSA2被定位于第5染色体SSR标记S5-57和S5-119之间,物理距离为718 kb。本研究为水稻遗传改良及研究温度影响叶绿体发育机制奠定了基础。     

Identification of restorers and maintainers for developing basmati and non-basmati hybrids in rice, Oryza sativa

Twenty‐seven improved aromatic lines of germplasm and 18 non‐aromatic disease‐resistant genotypes of rice were test‐crossed with four cytoplasmic male‐sterile lines (IR 58025A, IR 62829A, PMS 3A and PMS 10A). Thirteen aromatic and 10 non‐aromatic genotypes were selected based on pollen fertility, and crosses were repeated to confirm sterility‐maintaining and fertility‐restoring ability. Genotypes were categorized as effective restorers (> 80% spikelet fertility), partial restorers (21‐79% spikelet fertility) and maintainers (< 1% spikelet fertility). The effective basmati restorers identified were Basmati 385, Chandan, P1031‐8‐5‐1, HKR 241‐IET‐12020, SAF Khalsa 7 and Karnal Local. The basmati maintainers identified were Basmati 370, Pusa basmati 1, P615‐K‐167‐13 and P1173‐4‐1. The frequency of restorers obtained was higher for the non‐aromatic than the aromatic basmati type. The performance of restorers varied with cytoplasmic male‐sterile (CMS) line, location and season of testing. The differential ability to restore fertility in the CMS lines that have the wild abortive (WA) cytosterile system could result from different nuclear backgrounds of the CMS lines. These restorers and maintainers possess acceptable grain dimensions, a desirable degree of aroma, volume expansion through linear kernel elongation and cooking quality characteristics of basmati rice. These genotypes will contribute to developing basmati hybrids and provide restorers and maintainers with acceptable key basmati quality characteristics.     

利用SNP标记进行水稻品种籼粳鉴定

亚洲栽培稻(Oryza sativa L.)分为籼、粳2个亚种,随着杂交水稻的发展、种间杂种优势的利用,籼粳之间的界限变得越来越模糊。本研究利用3000份水稻种质资源信息,通过计算约2000万个单核苷酸多态性(single nucleotide polymorphism, SNP)位点的SNP-index值,进行籼粳特异SNP位点筛选,最终得到4084个籼粳特异SNP位点(4k-SNP);同时确定以籼粳指数作为水稻品种籼粳鉴定的指标。研究进一步采用大规模简单随机取样等统计分析方法对籼粳特异位点进行数据降维处理,将4k-SNP精简至40个SNP位点(40-SNP),用于水稻籼粳鉴定。为了验证40-SNP的籼粳鉴定效果,本研究一方面利用水稻生产上推广的82份选育品种,对40-SNP籼粳鉴定结果与4k-SNP鉴定结果进行比较,结果发现40-SNP与4k-SNP得出的粳型指数非常接近,相关系数为0.99;另一方面利用全球6类型(indica、aus、rayada、aromatic、tropicaljaponica、temperatejaponica)水稻品种共49份材料,对40-SNP籼粳鉴定...     

水稻半外卷叶突变体sol1的表型分析与基因定位

适度卷曲有利于提高水稻叶片的光合效率,增加植株光合产物的有效积累量。我们利用甲基磺酸乙酯(EMS)处理籼型水稻保持系西农1B,获得一个稳定遗传的水稻半外卷叶突变体。该突变体从十叶期开始各叶片逐渐向外卷曲直至半卷状,并伴随茎秆半矮化和叶片披垂,暂被命名为semi-outcurved leaf 1 (sol1)。与野生型(WT)相比, sol1的叶片卷曲指数均达到30%以上(P<0.01);倒一、倒二、倒三、倒四节节间长度和穗长极显著缩短,倒一、倒二、倒三叶的叶夹角显著或极显著增加;有效穗数、千粒重、每穗实粒数、结实率显著或极显著下降,一次枝梗数则增加11.3%(P<0.05)。sol1的蒸腾速率、胞间CO2浓度、气孔导度显著高于野生型。石蜡切片显示, sol1倒一叶的泡状细胞体积变小,数量显著增多,表皮细胞体积略微增大。遗传分析表明, sol1的半外卷叶性状受1对隐性核基因调控,定位于6号染色体标记JY6-3和JY6-10之间165kb的物理范围内,共含15个注释基因。qRT-PCR结果表明,与泡状细胞相关的内卷基因和外卷叶基因RL14、Roc5、REL1在突变体sol1中呈...     

High-resolution mapping revealed a 1.3-Mbp genomic inversion in Ssi1, a dominant semidwarf gene in rice (Oryza sativa)

Dwarf or semidwarf characters are an important trait for crop breeding as they provide lodging resistance. The rice mutant line DMF-1 has a dm-type semidwarf phenotype and high-lodging resistance controlled by the dominant gene, Short second internode 1 ( Ssi1 ). To elucidate the mechanism of reducing culm length in DMF-1, we sought to identify the Ssi1 gene by positional cloning using the chromosome segment substitution line as a crossing parent. As a result of high-resolution mapping, we found a 1.3-Mbp genomic inversion and a newly arranged gene in the Ssi1 locus. In this study, we report the high-resolution mapping and physical mapping of Ssi1 . We also discuss the possible function of a novel rearranged Ssi1 gene for the dominant dm-type semidwarf phenotype.