Waxy gene haplotypes: Associations with apparent amylose content and the effect by the environment in an international rice germplasm collection

Apparent amylose content (AAC), the key determinant of rice end-use quality attributes, is primarily controlled by the Waxy gene which codes for granule bound starch synthase (GBSS). We examined the combination of sequence variation in the Waxy gene and environmental effects, and their associations with AAC using 171 rice accessions originating from 43 countries. The combination of two single-nucleotide-polymorphism (SNP) markers in the Waxy gene allows for the identification of three marker haplotypes in this gene. The first SNP is at the leader intron splice site (In1 SNP), and the second polymorphism is in exon 6. The haplotypes explained 86.7% of the variation in AAC and discriminated the three market classes of low, intermediate and high AAC rice from each other. The environment affected the AAC of all haplotypes. Higher air temperature during grain development associated with a decrease in AAC of low and intermediate AAC-types, but with an increase in AAC of high AAC-type. The association of AAC with several Waxy RM190 microsatellite-(CT_n) alleles in combination with the In1 SNP was also examined. In conclusion, the Waxy haplotypes studied appear to be useful markers for selecting the AAC of breeding lines developed from the world's rice germplasm.     

Inheritance of flour paste viscosity is associated with a rice Waxy gene exon 10 SNP marker

Apparent amylose content is a key element for characterizing a rice (Oryza sativa L.) cultivar for cooking quality. However, cultivars with similar apparent amylose content can have widely varying quality attributes, including major parameters of flour paste viscosity. It has been postulated that the presence of a rice Waxy gene single nucleotide polymorphism (SNP) marker is associated with elevated Rapid Visco Analyser (RVA) properties in specific high amylose rice cultivars. A mapping population derived from a cross between two varieties, Cocodrie and Dixiebelle, having similar high apparent amylose contents, but with different paste viscosity properties and Waxy gene markers was analyzed for the genetic segregation of various pasting properties, measured with RVA instrumentation. Marker inheritance analyses revealed that the Waxy exon 10 SNP marker was associated with the proportion of soluble to insoluble apparent amylose and most RVA pasting measurements. Waxy gene markers can be used to efficiently improve the selection of rice with desirable characteristics, particularly for superior parboiling and canning quality.     

基于Waxy基因序列的西南地区玉米地方品种的演化

以40份西南地区玉米地方品种群体为材料,对Waxy基因第9外显子和第14外显子之间的DNA序列进行测序,将测序得到的40条序列与Gen Bank中来自普通玉米、小颖玉米、墨西哥玉米和二倍体多年生玉米的20条同源序列作比对分析。结果表明,来自西南地区四川、重庆、云南和贵州的玉米地方品种群体分别有180、152、163、164个多态性位点,供试种群遗传多态性指数依次为小颖玉米四川地方品种墨西哥玉米重庆地方品种二倍体多年生玉米贵州地方品种云南地方品种普通玉米。Tajima’D、Fu and Li’D和Fu and Li’F检验表明,Waxy基因是中性进化基因。比较种群间Waxy基因序列的共有多态性和固有差异,玉米地方品种与墨西哥玉米和二倍体多年生玉米间存在较高的固有差异和较低的共有多态性。Waxy基因的系统演化分析结果表明,小颖玉米、普通玉米和玉米地方品种有相近的遗传关系,小颖玉米是普通玉米和玉米地方品种的共同祖先。     

Development of new markers to genotype the functional SNPs of SSIIa, a gene responsible for gelatinization temperature of rice starch

Gelatinization temperature (GT) is an important quality predictor that determines the cooking quality of rice. GT is genetically controlled by the starch synthase IIa (SSIIa) gene. Two functional single nucleotide polymorphisms (SNPs) inside the SSIIa have already been found to be responsible for the variation of GT. One of these, GC/TT SNP at 4329/4330 bp, could be genotyped by four primers in a single PCR ( Bao et al., 2006a), but another one, G/A SNP at 4198 bp, has not been detected by a PCR-based marker. Here, we developed cleaved amplified polymorphic sequence (CAPS) and derived cleaved amplified polymorphic sequence (dCAPS) markers to detect these SNPs. A dCAPS marker that the PCR products were cleaved by the BseR I restriction endonuclease was designed to detect GC/TT SNP. Both CAPS and dCAPS markers were designed to detect G/A SNP using the restriction endonuclease Nla III and Tsp45 I, respectively. All the markers developed were co-dominant. It was known that the A allele of G/A SNP was rare among rice germplasm, but it was still in use by rice breeders. 11 rice accessions including landrace and breeding lines with A allele of G/A SNP were detected. The F₂ individuals from two crosses were used to analyze the co-segregation between the SNP alleles and the GT. The segregation ratio of two SNPs did not conform to the expected Mendelian ratio of 1:2:1, but the SNPs were co-segregated with GT. The markers developed in the present study would be useful in molecular breeding for the improvement of the quality of rice grain.     

Genetic relationship between grain chalkiness,protein content,and paste viscosity properties in a backcross inbred population of rice

A backcross inbred line population derived from a cross between Koshihikari and Kasalath was used to dissect the genetic relationship among chalkiness, protein content, and paste viscosity properties in rice in three environments. A total of 11 traits (or parameters) were analyzed, including percentage of grains with chalkiness (PGWC), protein content (PC) and protein index (PI), and eight parameters from the viscosity profile. PGWC, PC and PI were significantly correlated with the paste viscosity parameters. We identified 39 QTLs in three environments; ten QTL clusters emerged. Eight QTLs were consistently detected across the three environments and further confirmed using a set of chromosome segment substitution lines (CSSLs) where Kasalath was used as the donor parent and Koshihikari as the recurrent parent. One and two major clusters on chromosome 6 corresponded to the Wx and Alk loci, respectively. The former was responsible for PGWC and most of the viscosity parameters, and the latter for PI and some viscosity parameters. Particularly, QTL qPI-6.1 was linked with both the Wx and Alk loci. The co-locations of QTLs for PGWC and viscosity parameters and the linkage of qPI-6.1 and qBDV-6 at the Wx locus could be largely responsible for the phenotypic correlations between these traits.     

Investigating early vigour in upland rice (Oryza sativa L.): Part II. Identification of QTLs controlling early vigour under greenhouse and field conditions

Early vigour is an important characteristic for direct-seeded rice systems. The genetic control of early vigour was studied using a population of 129 backcross lines derived from a cross between Vandana, an improved indica, and Moroberekan, a traditional japonica. Screening was conducted under controlled conditions in greenhouse and field conditions, and indicators of early vigour, including shoot length, shoot biomass, leaf area, number of roots, root biomass, partitioning coefficients, and growth rates, were measured. Phenotypic correlations suggested that traits that were related and combined could be used to define early vigour. Broad-sense heritability ranged from moderate to high. Many regions were identified containing more than one QTL, suggesting that these traits were controlled by pleiotropic and/or closely linked QTLs. Many QTLs were specific to one environment but G × E interaction analysis showed that the main effects of the environment were large. Differences in temperature between experiments resulted in large differences in seedling age when expressed in thermal time. Different genes (QTLs) may be expected to control growth at different time intervals and thus may partly explain the limited agreement between experiments. However, several regions showed co-location of QTLs from more than one experiment. Comparisons with published studies revealed that these regions were previously identified in different genetic backgrounds and could potentially be used as introgression targets in a marker-assisted breeding program to improve germplasm for direct-seeded environments.     

高AC水稻淀粉合成相关酶基因表达的温度效应

以高直链淀粉含量(AC)早籼稻品种嘉育353为材料,利用人工气候箱设置高温(日均温32℃)和适温(日均温22℃)2个温度处理,采用实时荧光定量PCR技术分析了不同灌浆温度下水稻胚乳中3个淀粉分支酶基因(sbe1、sbe3和sbe4)、3个淀粉脱支酶基因(R酶基因、isa1和isa2)以及2个淀粉合酶(GBSSⅠ和SSSⅠ)基因的表达特性。结果显示,除了sbe4、isa1和GBSSⅠ基因外,其余基因高温下表达量均显著高于适温,表明sbe1、sbe3、R酶基因、isa2以及SSSⅠ基因属于温度敏感型基因,可能在水稻胚乳淀粉合成的温度调控方面起着关键作用。     

Response of lowland rice to agronomic management under different hydrological regimes in an inland valley of Ivory Coast

A large proportion of the rice in West Africa is produced in rainfed lowland ecosystems, mainly in inland valleys. The hydrological conditions (duration and intensity of flooding) vary with the toposequence position between the fringe and the centre of the valley. Production methods tend to evolve from the currently predominant unbunded plots without external input use, to input-intensive production in bunded plots. Agronomic management interventions co-evolve and may include varietal choice, herbicide use, and mineral N fertilizer application. The response of rice and the associated weeds to such interventions is likely to vary with the prevailing hydrological regime. A 2-year field experiment was conducted in northern Ivory Coast to determine the impact of water regime (plot position in the valley, presence of bunds) and input use (mineral N fertilizer and herbicide) on the productivity (yield and N use efficiency) of traditional and modern rainfed lowland rice cultivars and the biomass and composition of the associated weeds. Installing field bunds reduced seasonal variations in ponded water depth and resulted in a mean increase in rice grain yield of 30–40% (p < 0.005). This increase was associated with a 25% lower cumulative weed biomass and a several-fold increase in the agronomic use efficiency of applied mineral N in bunded than unbunded plots. Under low input management, traditional varieties tended to out-yield modern varieties in unbunded plots. Improved crop management such as herbicide and fertilizer application, and the construction of field bunds was more effective to increase the yield and N use efficiency in the flooded valley centre than in the drought-prone valley fringes. There is a need for site-specific targeting of modern cultivars, land development and improved production methods in the inland valleys of the West African savanna zone.     

Modeling the dependence of the crop calendar for rain-fed rice on precipitation in Northeast Thailand

Rain-fed lowlands are major agricultural ecosystems used for rice production in Northeast Thailand. Developing a tool to assess the effects of variable water conditions on the regional scale yield is also important to predict the effects of climate change on food supply. To estimate regional yields, we need a simple but accurate measure of the crop calendar (i.e., the distribution of planting dates), which has a strong influence on grain yield. In this article, we modeled the dependence of the crop calendar on rainfall patterns based on a survey of the region’s farmers as a part of an effort to provide a stronger basis for regional yield estimates. Our survey, conducted in 11 provinces for 2 years, confirmed the existence of large windows for sowing and transplanting versus narrow windows for heading and harvesting for rain-fed lowland rice culture in all the provinces. Variable water, soil, and toposequential conditions in the paddy fields were responsible for the large sowing and transplanting windows, whereas the use of photoperiod-sensitive varieties explained the narrow windows for heading and harvesting. The crop calendar was well expressed as a function of cumulative precipitation from June onward. When the crop calendar model was combined with a simple phenology-based model that uses growing degree-days adjusted by a day-length factor, we could estimate the rice crop calendar under rain-fed lowland conditions with acceptable accuracy. The model described in this article will be combined with a crop growth model to improve regional yield estimates for rain-fed lowland rice.     

Genetic relationship between grain yield and the contents of protein and fat in a recombinant inbred population of rice

To study the genetic relationship between grain yield and the nutrient contents in rice, 209 recombinant inbred lines derived from a cross between indica rice Xieqingzao B and Milyang 46 were used to determine quantitative trait loci (QTLs) affecting the yields and contents of the two major nutritional components in brown rice. Seven traits were analyzed, including brown rice recovery (BRR), protein content (PC), fat content (FC), grain yield (GYD), brown rice yield (BRYD), protein yield (PYD) and fat yield (FYD). The nutrient contents were significantly negatively correlated with BRR, GYD and BRYD, and the variations on nutrient yield were mainly ascribed to GYD. A total of 22 QTLs distributed on 10 regions of eight chromosomes were detected. Two QTL clusters were found on the short arm of chromosome 6 and the long arm of chromosome 10, respectively. The former was responsible for all the seven traits, and the latter for all the traits except BRR and FC. In both regions, the maternal alleles decreased nutrient contents, but they increased the yields of grain, brown rice, protein and fat. Implication of these results for the breeding of rice varieties with enhancing nutritional capacity is discussed.     

Grain quality characteristics of rice in Madagascar retail markets

A survey of rice in retail markets of Madagascar with emphasis on the surplus regions of Marovoay and Lac Alaotra and the deficient area, Antananarivo City showed rices of Madagascar to be predominantly medium-long, medium-shaped, with high apparent amylose content, low gelatinization temperature, and soft, hard or medium gel consistency. Mean protein content was 7.5%. Percent white grains among red and white grains ranged from 0 to 100% (mean 56%), head rice 0–94%, translucency 6–66% and Kett whiteness 12–48%.     

Identification of deletion mutants with improved performance under water-limited environments in rice (Oryza sativa L.)

Rice (Oryza sativa L.) is a semi-aquatic member of the grass family that is poorly adapted to dry environments and has greater sensitivity to water-deficits than other important cereals in this family. To increase productivity in aerobic or water-limited environments rice must overcome its adaptations to flooded environments. Deletion mutants offer an alternative genetic resource for improving drought tolerance. Almost 3500 IR64 deletion mutants were screened under vegetative and reproductive stage drought stress in the field and evaluated for leaf drying and/or grain yield. Seven novel conditional mutants of rice which showed gain of function through continued growth as drought stress developed compared to the wild type were identified. Mutant recovery rate was 0.1%. Further evaluation of putative drought mutants revealed that their average shoot biomass at maturity and grain yield per plant under stress exceeded those of the wild type by two-fold. Studies under controlled conditions confirmed mutants to have continued growth of both roots and shoots as drought developed compared to the wild type, and a tendency for greater water extraction. We propose that deletions in these mutants have affected a regulator of the highly conservative growth response common to irrigated lowland rice cultivars. Our results suggest that screening deletion mutants for performance under managed drought stress in the field could be a highly effective way to identify valuable genetic resources for improved drought response and aerobic adaptation in rice.     

Positional variations in phytic acid and protein content within a panicle of japonica rice

Six japonica rice cultivars differing in phytic acid and protein content were used to investigate the effect of grain position in the panicle on grain weight, phytic acid and protein content. There were significant differences in grain weight, albumin, globulin, prolamin, glutelin, and total protein content among the grains within a panicle for all six cultivars, but no differences were found for phytic acid content. An effect of grain position between primary rachises on grain weight and protein content was clearly detected, while that among top, middle and bottom rachis showed a greater change, varying with the cultivar. Albumin and globulin contents tended to be lower in the grains on the primary rachis than in those on the secondary rachis, and glutelin and total protein contents showed the opposite trend in the three cultivars with lower protein content. There was no significant correlation between phytic acid and protein content in the different grain positions within a panicle. The accumulation of phytic acid and protein during grain development appears to be independent.     

A model driven by crop water use and nitrogen supply for simulating changes in the regional yield of rain-fed lowland rice in Northeast Thailand

Climate change will have significant impacts on the rain-fed rice production ecosystem, and particularly on the ecosystem’s hydrology and water resources. Under rain-fed lowland conditions, substantial variations among fields in grain yield are commonly observed, but a method that can account for field-scale yield variability to produce regional-scale yield estimates is lacking, thereby limiting our ability to predict future rice production under changing climate and variable water resources. In this study, we developed a model for estimating regional yields of rain-fed lowland rice in Northeast Thailand, by combining a simple crop model with a crop calendar model. The crop model incorporates the effects of two important resources (water and nitrogen) on crop growth. The biomass accumulation is driven by water use, whereas the nitrogen supply determines canopy development and thereby constrains crop water use. Accounting for the wide range of planting dates and the strong photoperiod-sensitive characteristics of rice varieties through the calendar model is an essential component in determining regional yield estimates. The present model does not account for the effects of mid-season drought or flooding, but was nonetheless able to explain the spatial and temporal yield variations at the province level for the past 25 years. Thus, it can be used as a prototype for simulating regional yields of rain-fed lowland rice.     

The Null-4A Allele at the Waxy Locus in Durum Wheat affects Pasta Cooking Quality

Granule-bound starch synthase, also known as the waxy protein catalyses the synthesis of amylose in wheat endosperm starch. In durum wheats, the genes encoding GBSS are present at the two Wx loci on chromosome 7A and 4A (a segment of 7B that has been translocated). Several null Wx-B1 (missing GBSS protein from chromosome 4A) durum lines were produced from crosses with null-4A bread wheats backcrossed to durum wheats. Semolina milled from 4 normal and 7 null-4A durum wheat lines grown over two seasons (1999 and 2000) in South Australia were analysed for amylose content, starch pasting properties as measured by the Rapid Viscoanalyzer (RVA), swelling power and starch damage, protein content and electrophoretic protein analysis. Spaghetti was prepared with a micro-scale extruder and the cooked pasta evaluated for cooking loss, firmness, stickiness and water absorption. The null-4A lines had significantly lower (ca. 5%) amylose content, higher starch peak viscosities and semolina swelling power. The pasta derived from the null-4A lines had lower cooking loss and in 1999 was more adhesive than the non-waxy lines. Cooking loss was correlated with amylose content, peak starch viscosity, swelling power of semolina and cooked pasta adhesiveness. Semolina swelling power was highly correlated with RVA peak viscosity. Waxy durum wheats appear to have an advantage over the normal types in terms of lower cooking loss, widely used as an indicator of pasta cooking quality.     

A set of near-isogenic lines for blast resistance genes with an Indica-type rainfed lowland elite rice (Oryza sativa L.) genetic background

A set of near-isogenic lines for blast resistance genes was developed by using an Indica-type elite rice variety, IR49830-7-1-2-2, suitable for the rainfed lowland conditions in the tropics, as a genetic background. Initially, we revealed that IR49830-7-1-2-2 harbors five blast resistance genes - Pia, Pib, Pik-s, Pita, and Pi11(t) - by using a differential system involving 19 selected standard blast isolates from the Philippines. Based on this result, we developed nine near-isogenic lines (NILs) targeting eight resistance genes - Pik, Pi7(t), Pi3, Pi5, Pita-2, Piz-5, Pish, and Pi9 - by recurrent backcrossing. The introgression of each resistance gene in the NILs was confirmed by reaction patterns to the blast isolates, allelism tests, and DNA marker analysis. In addition, a genome-wide DNA marker survey revealed that most of the chromosome regions in each NIL were of the IR49830-7-1-2-2 type. The agricultural characteristics of most of the developed NILs were almost the same as those of IR49830-7-1-2-2. Moreover, with one exception, they showed submergence tolerance similar to IR49830-7-1-2-2. The developed NILs could be used as a multiline variety suitable for the rainfed lowland in the tropics.     

QTL mapping for crude protein and protein fraction contents in rice (Oryza sativa L.)

Improvement of rice storage proteins is important in rice breeding for high nutritional quality. Seventy-one recombinant inbred lines (RIL) derived from a cross between japonica variety Asominori and indica variety IR24 were used to study the inheritance of crude protein and protein fraction contents in rice. A total of 16 QTL were identified and mapped on eight chromosomes. Several QTL affecting contents of different protein fractions were mapped in the same chromosomal region. In particular, two QTL with a significant contribution were identified to simultaneously affect prolamin and glutelin contents. One QTL denoted as qCP-12 affecting crude protein content (CP) was located in the same region as QTL qGLT-12 affecting glutelin content, in agreement with the positive correlation between glutelin level and protein content. QTL with larger genetic effects were further confirmed using two sets of chromosome segment substitution lines (CSSL), where Asominori and IR24 were used as the recurrent parents. By QTL comparative analysis, two QTL for CP, three for globulin content and one for prolamin content were located in the vicinity of CP QTL previously identified in polished rice. Application of these results in rice breeding is discussed.     

基于生物信息学的小麦ALAD基因的鉴定和分析

5-氨基乙酰丙酸脱水酶(δ-aminoaevulinic acid dehydratase,ALAD)是生物体所有四吡咯化合物生物合成所必需的酶。为了给ALAD基因功能研究奠定基础,本研究根据模式植物拟南芥和水稻的ALAD基因序列,利用生物信息学手段对普通小麦ALAD基因进行鉴定和分析。结果表明,普通小麦有6个ALAD基因,其中TaALAD-A1、TaALAD-B1和TaALAD-D1基因位于第七同源群染色体长臂,而TaALAD-A2、TaALAD-B2和TaALAD-D2基因位于第六同源群染色体短臂。6个ALAD基因均含有保守的ALAD结构域,但系统发育分析中可分为2组。转录组数据表达分析结果显示,TaALAD-1基因在各个组织中的表达量均高于TaALAD-2,但两者在叶片中的表达量均最高。干旱胁迫对小麦TaALAD-1和TaALAD-2的表达几乎无影响;TaALAD-1的表达量在热胁迫1h下调,而在热胁迫6h时上调;热胁迫1h和6h对TaALAD-2的表达几乎无影响。通过小麦TILLING数据库分析,6个TaALAD基因都找到了数量不同的突变位点,其中一些突变位点为起始位点突变、无义突变、转录终止位点突变和剪切位点突变,可能影响该基因的功能。     

小麦条锈菌毒性变异与DNA多态性的相关性

为了解小麦条锈茵毒性和DNA多态性之间的相关性,为SSR标记技术在条锈茵群体遗传分析中的应用提供理论依据,对90个小麦条锈茵茵系进行了毒性和DNA多态性分析.毒性分析发现17种致病类型或生理小种,SSR分子标记分析发现75个基因型,DNA指纹的多态性远比毒性的多态性丰富.不同小种有相同或相近的DNA指纹,同一小种可能有不同的DNA指纹.毒性特征和DNA多态性之间并不存在相关性,SSR标记技术可用于小麦条锈茵群体遗传分析.     

Development of introgression lines of an Indica-type rice variety, IR64, for unique agronomic traits and detection of the responsible chromosomal regions

A total of 334 introgression lines (INLs: BC₃-derived lines) derived from crosses between a recurrent parent of Indica rice cultivar IR64 and 10 donor parents, including new plant type (NPT) lines IR65600-87-2-2-3, IR65598-112-2, IR65564-2-2-3, IR69093-41-2-3-2, IR69125-25-3-1-1, Hoshiaoba, IR66215-44-2-3, IR68522-10-2-2, IR71195-AC1, and IR66750-6-2-1, have been developed. These INLs with IR64 genetic background were characterized for eight agronomic traits: days to heading, culm length, leaf width, leaf length, panicle length, panicle number, 100-grain weight, and total spikelet number per panicle at the International Rice Research Institute from 2005 to 2007. To identify introgressed segments from the donor parents, genotypes of the 334 INLs were detected using more than 200 polymorphic simple sequence repeat markers. These segments detected on chromosomes 1, 2, 4, 5, and 6 were commonly introgressed across the INLs from more than four donor varieties. Based on the data of phenotype and genotype for the 334 INLs, associations between agronomic traits and introgressed chromosomal segments in the 334 INLs were investigated. A total of 54 regions for the eight traits were detected: seven regions for days to heading, eight regions for culm length, eight regions for leaf width, four regions for leaf length, six regions for panicle length, three regions for panicle number per plant, seven regions for 100-grain weight, and 11 regions for total spikelet number per panicle. Among them, the region on the long arm of chromosome 4 was associated with characteristics of the NPT such as long leaf, broad leaf, and high spikelet number. The developed 334 INLs with the IR64 genetic background will be useful materials for genetic analysis of agronomic traits.