直链淀粉含量不同的稻米淀粉结构、糊化特性研究

选用直链淀粉含量不同的4个水稻品种,研究其淀粉体结构和糊化特性。发现不同品种的淀粉体结构、糊化特性随直链淀粉含量的不同呈现一定的变化规律。直链淀粉含量从3.43%至21.65%,其晶体类型为A型,相对结晶度呈下降趋势,消减值表现为升高趋势,相对结晶度和消减值与直链淀粉含量分别呈显著负相关和极显著正相关(r=－0.964*, r=0.997**),消减值与结晶度呈显著负相关（r=－0.960*）。米粒的背部和腹部间淀粉体大小和排列有所不同,糯性或低直链淀粉含量与非糯性或高直链淀粉稻米的胚乳结构存在差异,淀粉体排列以糯稻较紧,粳稻次之,籼稻最松。淀粉的相对结晶度可能与胚乳中淀粉体的形状和紧密程度有关。     

Genetic studies of speciation in cultivated rice. 5. Inter- and intraspecific differentiation in the waxy gene expression of rice

Summary To get an insight in the gene regulation at the waxy locus of rice, the Wx gene product (Wx protein) controlling the synthesis of amylose was examined by electrophoretic techniques. Among nonwaxy rice strains, two different alleles, Wx ^a and Wx ^b, were found at the waxy locus. Wx ^a drastically enhanced the quantitative level of Wx protein as well as the amylose content in endosperm starch as compared with Wx ^b. The alleles acted additively in triploid endosperms. This implies that regulatory elements responsible for the Wx gene expression are on the same chromosome. The distribution patterns of Wx ^a and Wx ^b in five species of Oryza revealed that the regulatory changes are closely related to racial differentiation within a common rice species (O. sativa), suggesting that Wx ^b might have been selected for through the difference in grain quality during domestication.     

Isolation and characterization of a rice mutant insensitive to cool temperatures on amylose synthesis

The Wx ^b gene, one of the alleles at the rice waxy(wx) locus, is activated at cool temperatures during seed development, andas a result, a large amount of amylose is accumulated causing a reductionin rice grain quality. We found that the seeds of a du mutant couldbe visibly distinguished depending on whether they matured at cool ornormal temperatures. Using these characteristics, we isolated a mutantcandidate insensitive to cool temperatures. While the amylose content inthe original line was about 2% at a normal temperature (28 °C)and 12% at a cool temperature (21 °C), in the mutant candidate(coi) the amylose content was not affected by temperatures, i.e. theamylose content was about 3% at both temperatures. This finding incombination with the results of an immunoblot analysis indicated that theabsence of an increase in the amylose content in this mutant was caused bya constant level of Wx gene expression at normal and cooltemperature. Genetic analysis revealed that this insensitivity to cooltemperatures was caused by a single recessive mutation. This mutantshould be useful in breeding programs designed to produce rice of desiredquality at cool temperatures and in understanding genetic and molecularmechanisms that respond to slight changes in temperature.     

Characterization of waxy grain sorghum lines in relation to granule-bound starch synthase

The waxy phenotype, associated with endosperm containing little or no amylose, has been recognized in sorghum (Sorghum bicolor L. Moench) since 1933. Although variants of the waxy gene are well characterized in other cereals, the waxy trait has been assumed to be controlled by a single allele, wx, in sorghum. Recent improvements in technologies encourage re-examination of the waxy sorghums. The objectives of this research were therefore to identify and characterize sorghum lines with differing waxy alleles and to describe the actions of those alleles in crosses. Grain of eight waxy sorghum lines (BTxARG1, BTx630, Tx2907, B.9307, 94C274, 94C278, 94C289, 94C369), three wild-type checks (BWheatland, RTx430, BN122), and F₂ families from crosses among a subset of these lines were evaluated for presence or absence of granule-bound starch synthase (GBSS), the gene product of the wx locus, and wild-type vs. waxy endosperm. The F₂ segregation ratios were tested for fit to a 3:1 ratio using Chi-square analyses. Two distinctly different naturally occurring waxy alleles were identified: One with no GBSS (GBSS−), and one with apparently inactive GBSS present (GBSS+). We propose that the waxy allele with no GBSS be designated wx^a, and that waxy allele with apparently inactive GBSS present be designated wx^b. These two alleles are located in close proximity on the waxy locus. The wx^b allele is dominant to the wx^a allele in terms of GBSS production, and both are recessive to the wild-type Wx in terms of amylose content. The U.S. Government's right to retain a non-exclusive, royalty-free license in and to any copyright is acknowledged.     

QTL for rice grain quality based on a DH population derived from parents with similar apparent amylose content

A doubled haploid (DH)population consisting of 135 lines, derived from an indica (IR64) and a japonica (Azucena) rice with a similar apparent amylose content (AAC), was used to investigate the genetic factors affecting cooking and eating quality of rice. AAC,gelatinization temperature (GT), gel consistency (GC) and six starch pasting viscosity parameters were measured for quantitative trait loci (QTL) analysis using 193 molecular markers mapped on the DH population. A total of 17 QTLs were detected for the 9 traits, with at least one QTL and as many as 3 QTLs for each individual trait. No QTL for the measured parameters was found at the wx locus,possibly because of the similar AAC between the parents. Several QTLs with important effects on the variations in the measured parameters were detected in the present study which have not been found in earlier reports based on populations derived from parents with different AAC and wxgene alleles. Two interesting loci could be deduced from the present study according to the marker order compared with other genetic linkage maps. A QTL flanked by Amy2A and RG433 on the end of the long arm of chromosome 6, identified for GT, set back and consistency viscosity, might cover the gene encoding starch branching enzyme I. Similarly, a QTL flanked by RG139 and RZ58on chromosome 2, detected for hot paste viscosity and breakdown viscosity, might cover the gene encoding starch branching enzyme III. Generally, traits significantly correlated with each other shared identical QTL, but it was not true in some cases. The fine molecular mechanisms underlying these traits await further elucidation for the improvement of eating and cooking quality of rice. This revised version was published online in August 2006 with corrections to the Cover Date.     

Variation of <Emphasis Type="Italic">Wx</Emphasis> microsatellite allele,waxy allele distribution and differentiation of chloroplast DNA in a collection of Thai rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Eighty-two varieties of rice from different regions in Thailand were selected to explore the Waxy (Wx)gene diversity and indica-japonica differentiation of chloroplast DNA. A comparison of the 5 splice site in the first intron was made between glutinous and nonglutinous rice. It revealed that non-glutinous with low-amylose content and glutinous rice were characterized as the Wx^b allele based on the G-to-T base substitution, whereas non-glutinous rice with intermediate and high amylose carried the Wx^a allele. Four Wx microsatellite alleles, (CT)_n repeat, (n = 16,17,18 and 19) were found in glutinous rice. In contrast, non-glutinous rice showed five Wx microsatellite alleles (n = 11, 16, 17, 18 and 19). The (CT)₁₇ allele was prominent allele in Thai population, while the (CT)₁₁ allele was found only in intermediate and high amylose rice varieties from southern Thailand. Almost all of upland rice grown by various ethnic groups in northern Thailand were characterized as japonica type based on their having the PstI-12 fragment in their cpDNA, whereas most of rainfed lowland varieties from other regions of Thailand were indica. This exploration of DNA-based genetic markers is important, as it enhances our ability to describe and manipulate sources of genetic variation for rice breeding programs.     

Identification and verification of quantitative trait loci for eating and cooking quality of rice (Oryza sativa)

This study was conducted to identify new quantitative trait loci (QTLs) that have stable effects for eating and cooking quality (ECQ) of rice. Three recombinant inbred line populations of indica rice were each planted in two years. Three traits for ECQ, amylose content (AC), gel consistency (GC) and alkali spreading value (ASV), were measured for QTL analysis. A total of 13 QTLs were detected, including four for AC, six for ASV and three for GC. Two QTLs, qGC4 in the interval RM16252–RM335 on the short arm of chromosome 4 and qGC6.2 in the Alk region, were validated in a population derived from a residual heterozygote that was homozygous at the major locus Wx. In the absence of segregation at the Wx locus, qGC4 and qGC6.2 had additive effects of 2.46 and 8.18 mm, respectively, offering a potential for improving GC property of rice varieties. Comparison between qGC4 and previous results suggests that qGC4 is likely a new QTL for GC, providing a candidate for gene cloning and functional characterization.     

A gametocidal factor of Oryza glaberrima Steud. in O. sativa L.

Summary Recurrent backcrossing has been carried out with a view to transfer a gene for non-glutinous endosperm from two strains of O. glaberrima (Wx _g /Wx _g ) to glutinous japonica and indica varieties (wx/wx) of Oryza sativa. In the course of backcrosses Wx _g /wx segregants were crossed with each of the two glutinous varieties of sativa as the respective recurrent male parent. The wx/wx and Wx _g /wx segregants in the successive generations were consistently fully fertile and semi-sterile, respectively. The semi-sterility of Wx _g /wx plants was attributable to abortion of most of the pollen grains carrying the gene wx. The nucleus but not cytoplasm was related to the semi-sterility. The Wx _g /Wx plants having the gene for non-glutinous endosperm of a glaberrima strain and a japonica variety of sativa were also semi-sterile. Both wx- and Wx-megaspores in the plants heterozygous for the gene Wx _g were deleteriously affected. The results could be explained by assuming that a factor tightly linked with the gene Wx _g of glaberrima sterilizes gametes not carrying it in the heterozygotes and that the gametocidal action is exerted when combined with the sativa nucleus by the recurrent backcross method.     

Amylose synthesis capacity of the three Wx genes of wheat cv. Chinese Spring

The Wx locus controls amylose synthesis in the cereal endosperm. Hexaploid wheat (Triticum aestivum L.) has the three Wx loci on chromosomes 7A ( Wx-A1), 4A (Wx-B1) and 7D (Wx-D1). To verify the effects of null alleles on reducing amylose content and determine the amylose synthesis capacity of each Wx gene independently and accurately, we produced eight possible types of recombinant lines carrying different null alleles at the Wx loci under the ‘Chinese Spring’ genetic background. Amylose content varied from 0% of the waxy ‘Chinese Spring’ to 25% of the ‘Chinese Spring’ normal type. The reducing effect of the single null alleles was the largest in Wx-B1b, and there was no significant difference between Wx-A1b and Wx-D1b. More than 3% reductions in amylose content were detected in the double null types. The results of the double null lines further demonstrated that for the capacity of amylose synthesis, Wx-B1a predominates and produces 21–22% amylose, followed by Wx-D1a (20–21%) and Wx-A1a (15–18%). These significant differences were partly correlated with variation in the amounts of the Wx proteins produced by different Wx genes. However, comparisons of the double null lines with the single null or normal lines indicated that amylose content was not linearly proportional to the number of the Wx genes, suggesting that the Wx genes act in an epistatic manner. This revised version was published online in July 2006 with corrections to the Cover Date.     

Developing gene-tagged molecular markers for functional analysis of starch-synthesizing genes in rice (Oryza sativa L.)

The granule-bound starch synthase(GBSS), starch branching enzymes 1 (SBE1)and 3 (SBE3) are major enzymes involved in starch biosynthesis in rice endosperm. Available sequences of Sbe1 and Sbe3 genes encoding corresponding SBE1 and SBE3 have been used to identify homologous regions from genome databases of both the indica rice 93-11 and the japonica rice Nipponbare. Sequence diversities were exploited to develop gene-tagged markers to distinguish an indica allele from a japonica allele for both Sbe1 and Sbe3 loci. With these newly developed gene-tagged markers and available Wx gene markers, the roles of these starch-synthesizing genes (Sbe1, Sbe3, and Wx) in determining amylose content (AC) in the rice endosperm were evaluated using a double-haploid (DH)population derived from a cross between the indica rice cv. Nanjing11 and the japonica rice cv. Balilla. Only the Wx and Sbe3 loci had significant effects on the AC variation. On average, indica Wx ^a genotypes showed ∼9.1% higher AC than japonica Wx ^b genotypes, while indica Sbe3 ^a genotypes showed ∼1.0% lower AC than japonica Sbe3 ^b genotypes. A significant interaction was also observed between Wx and Sbe3 loci whereby the amylose content was 0.3% higher in Sbe3 ^a than Sbe3 ^b genotypes in the presence of the Wx ^a allele, but it was lower by 2.3% in the presence of the Wx ^b allele. Overall, polymorphisms at the Wx and Sbe3 loci together could account for 79.1% of the observed AC variation in the DH population. The use of gene-tagged markers in marker-assisted selection and gene functional analysis was also discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Development of near-isogenic lines of wheat carrying different null Wx alleles and their starch properties

The granule-bound starch synthase (GBSS I) encoded by the Wxgenes, is involved in amylose synthesis. For analyses of mechanisms of amylose synthesis and associated starch properties in hexaploid wheat, eight possible genotypes having different combinations of the three null alleles at the Wx loci with a common genetic background are a prerequisite. A near-isogenic population of doubled haploid (DH) lines was produced from Chinese Spring × waxy Chinese Spring F₁ plants using the wheat × maize method. The Wx protein phenotypes of the DH progeny were examined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and found that the null alleles at each of the three Wx loci segregated in a Mendelian fashion. A field trial demonstrated no differences between the eight types for ear emergence time, plant height and grain yield traits. Amylose content in the endosperm starch was highest in the wild type while lowest in the waxy type having no Wx proteins. Comparison between single null types and double null types indicated that the amylose synthesis capacity of Wx-A1a allele is the lowest. Pasting properties of starch are the highest in the waxy type, followed by the double null types. Consequently, both peak viscosity and breakdown were negatively correlated with amylose content. The chain-length distribution analysis of amylopectin structure revealed no clear difference among the eight types,suggesting that the reduced GBSS I activity due to introgression of the null Wx alleles does not affect either the chain length or the degree of branching of amylopectin. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effects of the two most common Wx alleles on different genetic backgrounds in rice

Wx^a and Wx^b are the most common alleles of the Wx gene in Asian cultivated rice. The difference between them is responsible for differences in the levels of the Wx gene product in the endosperm, as well as in amylose content, which is an important determinant of the quality of edible rice. Since Indica rice mostly carries the Wx^a allele, we introduced the Wx^b allele into Indica‐type rice (IR36) from a Japonica‐type rice (T65) by repeated backcrossing. In the near‐isogenic line (IR36 Wx^b), the level of the Wx gene product was reduced and, as expected, the level of amylose. However, IR36Wx^b had a lower amylose content than the recurrent parent of T65 with the Wx^b allele. The results suggest that the Indica (IR36) background might lower the amylose content more than the Japonica (T65) background when the Wx allele is the same. The possible importance of modifiers that regulate expression of the Wx gene is also discussed in relation to improvements in the grain quality of rice.     

Variation in cooking and eating quality traits in Japanese rice germplasm accessions

The eating quality of cooked rice is important and determines its market price and consumer acceptance. To comprehensively describe the variation of eating quality in 183 rice germplasm accessions, we evaluated 33 eating-quality traits including amylose and protein contents, pasting properties of rice flour, and texture of cooked rice grains. All eating-quality traits varied widely in the germplasm accessions. Principal-components analysis (PCA) revealed that allelic differences in the Wx gene explained the largest proportion of phenotypic variation of the eating-quality traits. In 146 accessions of non-glutinous temperate japonica rice, PCA revealed that protein content and surface texture of the cooked rice grains significantly explained phenotypic variations of the eating-quality traits. An allelic difference based on simple sequence repeats, which was located near a quantitative trait locus (QTL) on the short arm of chromosome 3, was associated with differences in the eating quality of non-glutinous temperate japonica rice. These results suggest that eating quality is controlled by genetic factors, including the Wx gene and the QTL on chromosome 3, in Japanese rice accessions. These genetic factors have been consciously selected for eating quality during rice breeding programs in Japan.     

Influence of single-nucleotide polymorphisms in the gene encoding granule-bound starch synthase I on amylose content in Vietnamese rice cultivars

Amylose content is one of the most important factors influencing the physical and chemical properties of starch in rice. Analysis of 352 Vietnamese rice cultivars revealed a wide range of variation in apparent amylose content and the expression level of granule-bound starch synthase. On the basis of single-nucleotide polymorphisms (SNP) at the splicing donor site of the first intron and in the coding region of the granule-bound starch synthase I gene, Waxy gene, alleles can be classified into seven groups that reflect differences in apparent amylose content. The very low and low apparent amylose content levels were tightly associated with a G to T in the first intron whereas intermediate and high amylose was associated with a T genotype at SNP in exon 10. The correlation between the combination of T genotype at SNP in the first intron, C in exon 6, or C in exon 10 was predominant among low amylose rice varieties. Our analysis confirmed the existence of Wx^op allele in Vietnamese rice germplasm. The results of this study suggest that the low amylose properties of Vietnamese local rice germplasm are attributable to spontaneous mutations at exons, and not at the splicing donor site.     

The effect of the Waxy locus (Granule Bound Starch Synthase) on pasting curve characteristics in specialty rices (Oryza sativa L.)

Starch structure and functionality have a significant impact on the utilization of cereal grains as food and feed. Starch viscosity characteristics are used to characterize rice cooking, processing and eating quality. In order to examine the genetics of viscosity characteristics, we developed molecular markers for five of the major enzymes involved in starch synthesis in the endosperm: granule bound starch synthase, soluble starch synthase, rice branching enzymes 1 and 3 and starch debranching enzyme. These markers were polymorphic in a cross between specialty rice varieties of diverging amylose content and viscosity characteristics. Our results indicate that the Waxy locus, encoding the gene for granule bound starch synthase, has a significant effect on peak viscosity, hot paste viscosity, cool paste viscosity, breakdown and setback viscosity. We estimate that the tightly linked (5–10 cm)locus for starch synthase may have a lesser, additive effect. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparison of Starch Levels Reduced by High Temperature During Ripening in Japonica Rice Lines Near‐Isogenic for the Wx Locus

One problem caused by high temperature during ripening in Japonica rice cultivars is a reduction in the amount of starch in the endosperm. To better understand this deleterious effect, we compared the accumulation of the two components of starch, amylose and amylopectin in grains ripened at high (32/28 °C) and low (22/18 °C) day/night temperatures in a set of lines of Japonica cultivar Taichung 65 (T65Wx^b) that are near‐isogenic for the Wx locus, which encodes granule‐bound starch synthase I. In T65Wx^b ripened at high temperature, the amount of starch per grain decreased. However, amylose per grain significantly decreased while amylopectin per grain significantly increased. On the other hand, the amount of amylopectin in T65wx, the amylose‐free line, did not differ significantly at the high and low temperatures. These data indicated that high temperatures during ripening did not directly affect amylopectin accumulation in T65Wx^b and that the reduction in starch in T65Wx^b from the high temperatures was caused by a decrease only of amylose. The results for T65Wx^a and T65Wx^op were also consistent with this conclusion. As a result of the decrease in amylose, the outer region of starch granules from T65Wx^b ripened at the high temperatures also had less I₂KI staining. Because this fact might suggest that a portion of amylose was synthesized inside the developing granules after amylopectin synthesis in rice, the effect of amylose deposition in increasing of the density of starch granules is also discussed.     

Amylose content and starch properties generated by five variant <Emphasis Type="Italic">Wx</Emphasis> alleles for granule-bound starch synthase in common wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

In common and durum wheats (Triticum aestivum L. and T. durum Desf.), variant waxy (Wx) alleles have been reported for three Wx proteins (Wx-A1, -B1 and -D1), responsible for amylose synthesis in flour starch. Five variant alleles, Wx-A1c, -A1e, -B1c, -B1d and -D1c, were examined to elucidate their effects on amylose content in flour starch. Common wheat lines carrying a Wx protein produced by one variant (e.g., Wx-A1c) and one control (e.g., Wx-A1a) allele were bred and their starches were compared. Results showed that Wx-A1e did not produce amylose (waxy phenotype), whereas three alleles (Wx-A1c, -B1c and -B1d) reduced amylose, and -D1c might have increased it slightly. Most data on blue value, swelling power and starch paste clarity in water and dimethyl sulphoxide also suggested the variant Wx alleles either reduced or increased amylose content.     

Gene dosage effect of the wheat Wx alleles and their interaction on amylose synthesis in the endosperm

To find out gene dose effect of each of the three homoeologous Wx genes and their interaction on the production of granule-bound starch synthase (GBSS I) and amylose biosynthesis in the endosperm, Chinese Spring and its near-isogenic waxy types were crossed reciprocally and, obtained a plant population with varying doses of each Wx gene. The amount of GBSSI was increased linearly with increasing gene dose of either of Wxloci. In each of the three Wx loci, the change in amylose content was linear up to 3 doses, with a more potent capacity ofWx-B1a at any dose. Higher level of amylose production was observed in the reciprocal F₁ grains than the expected effect of dose/s of each gene or additive effect of different allelic combination by artificially blend starches which have amylose produced by equivalent number ofWx alleles to that of relevant F₁ cross. When Wx-B1a and Wx-A1a were combined, increase in amylase content was not in proportion to increase in gene dosage. The enhanced amylase synthesis was shown by 2-gene and 3-geneinteraction, indicating that not only type of the three Wx genes and its dose but the interaction among them have significant roles in determining the amylose content. This revised version was published online in July 2006 with corrections to the Cover Date.     

Localization of QTL for basal root thickness in <Emphasis Type="Italic">japonica</Emphasis> rice and effect of marker-assisted selection for a major QTL

Root traits are key components of plant adaptation to drought environment. By using a 120 recombined inbred lines (RILs) rice population derived from a cross between IRAT109, a japonica upland rice cultivar and Yuefu, a japonica lowland rice cultivar, a complete genetic linkage map with 201 molecular markers covering 1,833.8 cM was constructed and quantitative trait loci (QTLs) associated with basal root thickness (BRT) were identified. A major QTL, conferring thicker BRT, located on chromosome 4, designated brt4, explained phenotypic variance of 20.6%, was selected as target QTL to study the effects of marker-assisted selection (MAS) using two early segregating populations derived from crosses between IRAT109 and two lowland rice cultivars. The results showed that the flanking markers of brt4 were genetically stable in populations with different genetic backgrounds. In the two populations under upland conditions, the difference between the means of BRT of plants carrying positive and negative favorable alleles at brt4 flanking markers loci was significant. Phenotypic effects of BRT QTL brt4 were 5.05–8.16%. When selected plants for two generations were planted at Beijing and Hainan locations under upland conditions, MAS effects for BRT QTL brt4 were 4.56–18.56% and 15.46–26.52% respectively. The means of BRT for the homozygous plants were greater than that of heterozygous plants. This major QTL might be useful for rice drought tolerance breeding. L. Liu and P. Mu are contributed equally to this work.     

Altered tissue-specific expression at the Wx gene of the opaque mutants in rice

Amylose content is a major determinant of the eating quality in rice. To elucidate the allelic diversity at the Waxy (Wx) gene which controls the amylose synthesis, two cultivated strains having opaque endosperms were studied. The gene responsible for opaque endosperms was introduced into the genetic background of the Japonica type of rice by successive backcrosses, and the two near-isogenic lines (NILs) were selected from the B5 generation. The genetic experiments revealed that an allele, Wx^op, controls opaque endosperms which show chalky as wx endosperms in spite of the production of amylose. Immunoblotting analysis was carried out to compare the gene expression by using the NILs with 4 different alleles (Wx^a, Wx^b, Wx^op and wx). The level of the gene product bound to starch granules was slightly lower in the NILs with Wx^op than that with Wx^b, showing a positive correlation with amylose content in the endosperm. Extracts from mature anthers indicated that the gene product was markedly reduced in the NILs with Wx^op as well as that with wx, showing an altered expression in the tissue specificity in the Wxop lines. Sequence analysis suggested that the Wx^op had been derived from Wx^a, independently of the origin of Wx^b. The importance of the gene regulation was discussed in relation to diversified phenotypes established during the domestication process.