Wx Locus Research Articles

Physicochemical Characteristics of Starches of Two Sets of Near‐isogenic Wheat Lines with Different Amylose Content

AbstractWheat lines with modified amylose content were produced by controlling the null alleles of three homoeologous Wx loci, Wx‐A1, Wx‐B1 and Wx‐D1 and those with triple‐null Wx alleles were found to yield waxy, amylose‐free starch. In this study, the new near‐isogenic lines developed by the recurrent backcross method between cv. Kanto 107 and the waxy mutant lines, K107Wx1 or K107Wx2, were cultivated and the physicochemical characteristics of their starches were investigated. The apparent amylose contents of the waxy near‐isogenic lines were 1.6 – 3.8%, whereas those of the non‐waxy near‐isogenic lines were 23.2 – 25.4%. The waxy lines had higher gelatinization temperature and enthalpy than the non‐waxy lines and no peak of amylose‐lipid complex as determined by DSC. The X‐ray diffraction patterns from both waxy and non‐waxy near‐isogenic lines corresponded to A‐type crystallization, like their parent lines. The starches isolated from the waxy lines had lower pasting onset and peak temperatures, higher peak viscosities and breakdown and lower final viscosity than those of the non‐waxy lines in the Viscoamylograph. The starch of non‐waxy near‐isogenic lines had lower peak and final viscosities and higher breakdown than did the recurrent parent line, cv. Kanto 107. The characteristics of the new wheat starches were well understood and classified, which will contribute to the wide application of these starches in food processing.

Genetic basis of 17 traits and viscosity parameters characterizing the eating and cooking quality of rice grain

A recombinant inbred line population derived from a cross between Zhenshan 97 and Delong 208 was used to analyze the genetic basis of the cooking and eating quality of rice as reflected by 17 traits (or parameters). These traits include amylose content (AC), gel consistency (GC), alkali spreading value (ASV), cooked rice elongation (CRE), and 13 parameters from the viscosity profile. All the traits, except peak paste viscosity (PKV), time needed from gelatinization to peak (BAtime), and CRE, can be divided into two classes according to their interrelationship. The first class consists of AC, GC, and most of the paste viscosity parameters that form a major determinant of eating quality. The second class includes ASV, pasting temperature (Atemp) and pasting time (Atime), which characterize cooking process. We identified 26 QTL (quantitative trait locus or loci) in 2 years; nine QTL clusters emerged. The two major clusters, which correspond to the Wx and Alk loci, control the traits in the first and second classes, respectively. Some QTL are co-located for the traits belonging to the same class and also for the traits to a different class. The Wx locus also affects on ASV while the Alk locus also makes minor contributions to GC and some paste viscosity parameters. The QTL clusters on other chromosomes are similar to the Wx locus or Alk locus, although the variations they explained are relatively minor. QTL for CRE and PKV are dispersed and independent of the Wx locus. Low paste viscosity corresponds to low AC and soft gel, which represents good eating quality for most Chinese consumers; high ASV and low Atemp, together with reduced time to gelatinization and PKV, indicate preferred cooking quality. The genetic basis of Atemp, Atime, BAtime, peak temperature, peak time, paste viscosity at 95 degrees C, and final paste viscosity is newly examined to reveal a complete and dynamic viscosity profile.

The Source of Genes Related to Rice Grain Starch Synthesis Among Cultivated Varieties and Its Contribution to Quality

The property of starch in rice grain endosperm is a very important determinant for rice quality, and it is essential to understand the genetic effect of the genes related to starch synthesis in high-yielding rice varieties for rice quality improvement. The physicochemical properties (e.g., amylose content, gel consistency, and RVA profile) were assessed on 53 rice varieties, including certain typical indica/japonica landraces and certain high-yielding modern varieties. And molecular markers for Sbel, Sbe3 developed on the basis of sequence diversities between the rice subspecies indica and japonica, together with PCR- Acc I marker for Wx gene were used to investigate the genotypes of 53 rice cultivars. The result showed that the developed molecular markers for Wx, Sbel, Sbe3 could distinguish indica or japonica alleles at three loci. Among all the 53 rice cultivars, six genotypes were observed when Sbel, Sbe3, and Wx loci were considered together, while the genotypes of Wx i Sbel j Sbe3 i and Wx i Sbel j Sbe3 i were absent. In order to explore the genetic effects of the three genes, especially for starch branching enzyme genes, ANOVA and multiple comparison analysis were conducted. The results showed that rice cultivars with different genotypes exhibited different phenotypes, including amylose content, gel consistency and certain RVA characteristics, and the significant differences among the six genotypes were observed. It was concluded that these three genes had randomly recombined during the process of the rice variety development. And the genetic effects of indica and japonica alleles at three gene loci were different, of which, Wx gene plays a major role in determining the starch properties, followed by Sbel and Sbe3, and the genetic effects of Sbel and Sbe3 in different backgrounds ( Wx i, Wx j) are different. The results have provided a clue for rice good quality variety development, and the molecular markers will benefit to the improvement in quality of rice.

Induction of a Large Deletion Including the waxy Locus by Thermal Neutron Irradiation in Rice

Sequence rearrangement of rice waxy (wx) mutant N26 induced by thermal neutron irradiation was analyzed by PCR amplification and subsequent DNA sequencing. We found that the genome region of Norin 8, including the Wx locus, had been deleted, corresponding to 39 867 nucleotides for the japonica cultivar Nipponbare. Sequence analysis suggested that a short homologous sequence was used for rejoining without any addition of nucleotides. This is the first mutant in which a complete loss of the Wx locus was induced by thermal neutron irradiation in rice.

Functional analysis of starch-synthesis genes in determining rice eating and cooking qualities

Apparent amylose content (AAC), gel consistency (GC), and gelatinization temperature (GT) are recognized as the most important determinants of rice eating and cooking qualities. The contributions of major starch-synthesis genes to these three traits have been investigated in the three consecutive experiments. In an initial QTL mapping with 130 doubled haploid (DH) lines, derived from an inter-subspecific cross of ‘Nanjing11’/‘Balilla’, the major QTLs responsible for AAC, GC, and GT coincided with the Wx (granule-bound starch synthase gene), Wx, and Sss IIa (soluble starch synthase gene) loci, respectively. In the second experiment, contributions of the major starch-synthesis genes to AAC, GC, and GT variations were estimated by using a multiple linear regression analysis. As shown, the Wx locus was a principal determinant for both AAC and GC, and could account for 58.5% and 38.9% of the phenotypic variations, respectively; while the Sss IIa locus was associated with GT, and could explain 25.5% of the observed variation. Eventually, a F2 population consisting of 501 individuals, derived from an inter-subspecific cross of the two sticky rice varieties ‘Suyunuo’ and ‘Yangfunuo 4’, was examined with gene-tagged markers. In the absence of the Wx gene, none of the starch-synthesis genes investigated could dominate the GC variation, however, the Sss IIa locus could also explain 25.1% of the GT variation. In summary, the Wx locus dominates the AAC variation, and meanwhile plays a major role in the GC variation. The Sss IIa locus is a major factor in explaining the GT variation. Apart from the major genes, other genetic factors may also contribute to the GC/GT variations.

Microsatellites, single nucleotide polymorphisms and a sequence tagged site in starch-synthesizing genes in relation to starch physicochemical properties in nonwaxy rice (Oryza sativa L.)

Starch characteristics determine the quality of various products of rice, e.g., eating, cooking and processing qualities. Our previous study indicated that molecular markers inside or close to starch synthesizing genes can differentiate the starch properties of 56 waxy rices. Here we report microsatellite (or simple sequence repeat, SSR) polymorphism in the Waxy (Wx) gene, soluble starch synthase I gene (SS1) and starch branching enzyme 1 gene (SBE1), single nucleotide polymorphism (SNP) in Wx and starch branching enzyme 3 gene (SBE3), and a sequence tagged site (STS) in starch branching enzyme 1 gene (SBE1) among 499 nonwaxy rice samples and their relationships with starch physicochemical properties. The nonwaxy rice samples consist of landraces (n = 172) obtained from germplasm centers and cultivars and breeding lines (n = 327) obtained from various breeding programs. Ten (CT)( n ) microsatellite alleles, (CT)(8), (CT)(10), (CT)(11), (CT)(12, )(CT)(17), (CT)(18), (CT)(19), (CT)(20), (CT)(21), and (CT)(22), were found at the Wx locus, of which (CT)(11) was the most frequent, and (CT)(12), (CT)(21) and (CT)(22) were identified for the first time. Four (CT)( n ) microsatellite alleles were found at the SBE1 locus, (CT)(8), (CT)(9), and (CT)(10 )together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion, of which (CT)(9) and the insertion was a new allele identified in only one rice, IR1552. Multiple microsatellites clustered at the SS1 locus, and in addition to the three alleles previously detected (SSS-A = (AC)(2)...TCC(TC)(11)...(TC)(5)C(ACC)(11), SSS-B = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(9), and SSS-C = (AC)(3)...TCT(TC)(6)...(TC)(4)C(ACC)(8)), one new allele (SSS-D = (AC)(2)...TCC(TC)(10)...(TC)(4)C(ACC)(9)) was found. Analysis of the starch physicochemical properties of the samples with different microsatellites, SNPs and STS groups indicated that these molecular markers can differentiate almost all the physicochemical properties examined, e.g., apparent amylose content (AAC), pasting viscosity characteristics, and gel textural properties. Wx SSR and Wx SNP alone explained more variations for all physicochemical properties than the other molecular markers. The total six markers could explain 92.2, 81 and 86% of total variation of AAC, gel hardness (HD), and gel cohesiveness (COH), respectively, and they could explain more than 40% of the total variation of hot paste viscosity (HPV), cool paste viscosity (CPV), breakdown viscosity (BD), setback viscosity (SB) and gel adhesiveness (ADH). However, only 29% of the total variation of peak viscosity (PV) and 37% of pasting temperature (PT) could be explained by all the molecular markers. Some of these markers can differentiate the starch physicochemical properties among the rice samples with the same Wx allele, indicating that the variation within Wx allele classes can be explained by other starch synthesizing genes. These SSRs, SNPs and STS are useful in marker-assisted breeding for the improvement of starch quality of rice.

Genetic Analysis of Long Chain Synthesis in Rice Amylopectin

The amount of long chains (LC) of amylopectin in high-amylose rice is thought to be one of the important determinants of its quality when cooked. A wide range of differences in LC content have been reported in rice varieties, which can be clearly divided into four classes based on LC and apparent amylose content: namely, amylose and LC-free, low or medium-amylose and low-LC, high-amylose and medium-LC, high-amylose and high-LC. However, genetic factors controlling LC content have not been fully understood. Here, we performed quantitative trait loci (QTL) analysis of LC content using 157 recombinant inbred lines (RILs) derived from a cross of a low-LC cultivar, Hyogokitanishiki, and a high-LC line, Hokuriku 142. By analyzing randomly selected 15 RILs, it was shown that high LC content (≥11%) was associated with high setback viscosity (≥200 RVU), and that low LC (≤ 3%) was associated with low setback viscosity (≤ 130 RVU), as measured by a Rapid Visco Analyzer. With setback viscosity as an indicator for LC content, QTL analysis was conducted using 60 DNA markers including a CAPS marker that distinguished Wx a and Wx b alleles coding for granule-bound starch synthase I (GBSSI or Wx protein), the enzyme working for amylose biosynthesis. Only one QTL with a peak log of likelihood score at the wx locus was detected, and no line showing setback viscosity corresponding to the medium-LC class appeared. The fact that wx mutants of Hokuriku 142 lacked LC in their rice starch supports the view that the functional Wx allele is indispensable for LC synthesis in addition to amylose synthesis in rice endosperm. We suggest three possible reasons why no line with medium-LC content was observed. First, the locus (loci) responsible for generation of medium-LC may be located very close to the wx locus and not able to be dissected by the population and DNA markers we used. Second, there may be special QTLs for medium-LC cultivars that do not exist in low- or high-LC cultivars. Third, medium-LC cultivars may have an as-yet unidentified Wx allele with lower capability in LC synthesis compared to the Wx allele in high-LC cultivars.

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 main effects, epistatic effects and environmental interactions of QTLs on the cooking and eating quality of rice in a doubled-haploid line population

Amylose content (AC), gel consistency (GC) and gelatinazation temperature (GT) are three important traits that influence the cooking and eating quality of rice. The objective of this study was to characterize the genetic components, including main-effect quantitative trait loci (QTLs), epistatic QTLs and QTL-by-environment interactions (QEs), that are involved in the control of these three traits. A population of doubled haploid (DH) lines derived from a cross between two indica varieties Zhenshan 97 and H94 was used, and data were collected from a field experiment conducted in two different environments. A genetic linkage map consisting of 218 simple sequence repeat (SSR) loci was constructed, and QTL analysis performed using QTLMAPPER 1.6: resolved the genetic components into main-effect QTLs, epistatic QTLs and QEs. The analysis detected a total of 12 main-effect QTLs for the three traits, with a QTL corresponding to the Wx locus showing a major effect on AC and GC, and a QTL corresponding to the Alk locus having a major effect on GT. Ten digenic interactions involving 19 loci were detected for the three traits, and six main-effect QTLs and two pairs of epistatic QTLs were involved in QEs. While the main-effect QTLs, especially the ones corresponding to known major loci, apparently played predominant roles in the genetic basis of the traits, under certain conditions epistatic effects and QEs also played important roles in controlling the traits. The implications of the findings for rice quality improvement are discussed.

Assessing the relationship between endosperm type and grain yield potential in sorghum ( Sorghum bicolor L. Moench).

Sorghum hybrids with waxy endosperm (either homozygous or heterozygous waxy) have lower grain yield potential compared to non-waxy endosperm hybrids. The cause of this yield reduction is not known. From a genetic perspective, the yield reduction could be due to pleiotrophy or genetic linkage between the wx locus and other loci that influence grain yield. The specific cause of the relationship is important because an effective breeding program can alleviate the problem if it is due to linkage. The objective of this study was to determine whether linkage or pleiotrophy is causing the negative relationship between grain yield and waxy endosperm. From each of two F 2 breeding populations segregating for waxy endosperm, between 40 and 50 inbred lines were derived, with equal numbers of waxy and non-waxy endosperm lines. No selection for yield was practiced during the development of these lines. The lines from these two populations and a set of testcross hybrids (derived from one population) were evaluated in four environments in Texas from 1998 to 2000. Across all tests and environments, the combined yield of the waxy genotypes was 17% lower than non-waxy genotypes. While yields were lower in waxy genotypes, analysis of the individual inbred lines and hybrids revealed that several waxy inbred lines were not statistically different from the best non-waxy inbreds. These results imply that selection of high yielding waxy genotypes is possible, but a significant breeding emphasis on their development is required to effectively identify those genotypes.

Reduced Amylose Effects on Bread and White Salted Noodle Quality

ABSTRACTAmylose content in wheat endosperm is controlled by three Wx loci, and the proportion of amylose decreases with successive accumulation of Wx null alleles at the three loci. The proportion of amylose is believed to influence end‐use quality of bread and Asian noodles. The objectives of this study were to determine influence of the allelic difference at Wx‐B1 locus on bread quality, bread firmness, and white salted noodle texture in a spring wheat cross segregating for the Wx‐B1 locus and in a set of advanced spring wheat breeding lines differing in allelic state at the Wx‐ B1 locus. In addition, we examined the relationship between amylose content and flour swelling properties on bread and noodle traits. Fifty‐four recombinant inbred lines of hard white spring wheat plus parents were grown in replicated trials in two years, and 31 cultivars and breeding lines of hard spring wheat were grown in two locations. Bread and white salted noodles were processed from these trials. The presence of the Wx‐B1 null allele reduced amylose content by 2.4% in a recombinant inbred population and 4.3% in a survey of advanced breeding lines and cultivars compared with the normal. The reduced amylose was accompanied by an average increase in flour swelling power (FSP) for the Wx‐B1 null group of 0.8 g/g for the cross progeny and 2.3 g/g for the cultivar survey group. The Wx‐B1 allelic difference did not affect flour protein in cross progeny where the allelic difference was not confounded with genetic background. Bread from the Wx‐B1 null groups on average had increased loaf volume and was softer than the normal group for the cross progeny and cultivar survey group. The Wx‐B1 allelic difference altered white salted noodle texture, most notably noodle springiness and cohesiveness where the Wx‐B1 null groups was more springy and more cohesive than the normal groups for both sets of genetic materials. Flour protein was more highly related to loaf volume than were FSP or amylose. Both flour protein and FSP were positively related to noodle textural traits, but especially noodle springiness and cohesiveness.

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.

Genes coding for starch branching enzymes are major contributors to starch viscosity characteristics in waxy rice (Oryza sativa L.)

Granule-bound starch synthase (GBSS) encoded by the Waxy (Wx) gene determines the synthesis of amylose; while, starch branching enzymes encoded by Sbe genes are involved in the formation of amylopectin. Both amylose and amylopectin form starch in the endosperm of seeds. Here, we report on the effects of the wx gene and the two Sbe genes, Sbe1 and Sbe3, on starch paste viscosity in waxy rice. Molecular markers for both Sbe1 and Sbe3 have been developed based on sequence diversity between the rice subspecies indica and japonica. All Sbe- and Wx- linked markers have been used to investigate 40 waxy rice cultivars. For the wx locus (null Wx locus in waxy rice), four classes of (CT)n repeats, including (CT)16, (CT)17, (CT)18, and (CT)19, were detected in the putative leader sequence. Moreover, a unique sequence, AGTTATA, is also present at the 5′ splice junction site of the putative leader intron. Overall, the wx locus has no effect on starch paste viscosity characteristics. Among all 40 waxy rice cultivars, five genotypes are observed at the Sbe1 and Sbe3 loci. Polymorphisms in both Sbe1 and Sbe3 loci account for ∼70% of the observed variations in both hot and cool viscosities, and for 40% of the observed variations in both peak viscosity and consistency.

Distribution of p-SINE1-r2 at the Wx locus in cultivated and wild rice in Thailand

Short interspersed elements (SINEs) are retrotransposons that make their copies by the retroposition of RNA made by RNA polymerase III (Motohashi et al 1997). These are generally found in animals and plants. Umeda et al (1991) and Mochizuki et al (1993) reported the existence of the p-SINE1 family in rice. Furthermore, Hirano et al (1994) reported that the p-SINE-r2 , a member of the SINE family, is found only in two closely related species with the AA genome , Oryza sativa and O.rufipogon . This element is located in the 10th intron of the Waxy gene (Umeda et al 1991). Mochizuki et al (1993) suggested that p-SINE1-r2 could be useful for classifying various rice strains with the AA genome.

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.

Isolation and Characterization of Starch Mutants in Rice

We isolated various kinds of mutants participating in the biosynthesis of endosperm starch by the MNU treatment of fertilized egg cells in japonica rice cultivars. There were at least 6 mutant loci lowering amylose content including the wx locus. These low amylose mutations altered the amylose content without affecting the amylopectin structure. The amylose content did not affect the initiation of gelatinization, but influenced greatly the swelling of gel, indicating that amylose content affects the swelling power of gel and the termination of gelatinization. BEIIb mutation decreased specifically the short chains with DP less than 17 in amylopectin whereas BEI mutation was characterized by a significant decrease in long chains with DP longer than 38 and short chains with DP 12 to 23, suggesting that BEIIb and BEI contributes to the synthesis of A chains, and B, and B3 chains of amylopectin, respectively. Notable alteration on the chain length profile was not observed in BEIIa mutation. Each BE mutation showed distinct effect on the thermal properties of endosperm starch. The sug2 mutation lowering the expression level of PUL increased shorter chains of amylopectin and accumulated a significant amount of the water-soluble polysaccharides as well as ISA (sugl ) mutation. The fl o2 mutation decreased simultaneously the expression levels of some of the enzymes including BEI, PUL and GBSS, suggesting that the wild type Flo2 gene probably encodes a regulatory protein that modulates the expression of the genes involved in starch biosynthesis. Some mutants involved in the biosyn thesis of ADP-glucose were also isolated.

Microsatellites in starch-synthesizing genes in relation to starch physicochemical properties in waxy rice ( Oryza sativa L.).

Rice starch is composed of amylose and amylopectin. Amylose content, an important determinant of rice starch quality, is primarily controlled by the waxy gene, encoding granule-bound starch synthase (GBSS). The starch branching enzyme (SBE) and soluble starch synthase (SSS) play major roles in the synthesis of amylopectin. Microsatellite polymorphisms in the three genes, the wx gene encoding granule-bound starch synthase I, the SBE gene encoding starch branching enzyme I and the SSS gene encoding soluble starch synthase I, were studied for 56 accessions of waxy rice ( Oryza sativa L.). Four (CT)(n) microsatellite alleles, (CT)(16), (CT)(17), (CT)(18) and (CT)(19), at the wx locus were detected in this set of waxy rice, of which (CT)(17) was the most frequent. Three (CT)(n) microsatellite allele classes were found at the SBE locus, (CT)(8) or (CT)(10) together with an insertion sequence of CTCTCGGGCGA, and (CT)(8) alone without the insertion. There were multiple microsatellites clustered at the SSS locus. However, these alleles can also be grouped into three classes, i.e. the allele class SSS-A = (AC)(2) em leader TCC(TC)(11) em leader (TC)(5)C(ACC)(11), the allele class SSS-B = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(9), and the allele class SSS-C = (AC)(3) em leader TCT(TC)(6) em leader (TC)(4)C(ACC)(8). The analyses of starch physicochemical properties among different microsatellite genotypes indicated that the waxy rice group with the (CT)(19) allele, the SBE-A allele and the SSS-B allele was quite different from other groups. Nine out of 15 accessions with a high gelatinization temperature (GT) belonged to the wx (CT)(19) group, all of them belonged to the SBE-A group and 13 of them belonged to the SSS-B group. These microsatellites might be useful in marker-assisted breeding for the improvement of rice grain quality.

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.

STS and microsatellite marker-assisted selection for bacterial blight resistance and waxy genes in rice, Oryza sativa L.

DNA marker-assisted selection was employed to select Xa-21 bacterial blight resistance and waxy (Wx) genes. Genotypes with both genes were selected from four F2 populations involving indica × indica, indica × intermediate and japonica × indica crosses. With the assistance of PCR marker, 13 true breeding lines carrying Xa-21 were identified from F2 generation of IRBB 21 × G 11353 cross. Similarly ten, eleven and fifty two plants having Xa-21 gene were isolated from G 3005-4-1 × IRBB 21, IRBB 21 × HJX 74 and IRBB 21 × SY 2crosses respectively. The lines with Wx gene in both homozygous and heterozygous state were also scored from the above crosses. Twenty plants having both Xa-21 and Wx loci in homozygous state were identified. DNA-based progeny testing was conducted to ensure the selection of homozygous lines for Xa-21 and Wx genes. Finally,twenty true breeding lines with high amylose content and Xa-21 gene were isolated from four crosses. These homozygous lines are phenotypically superior and resistant to Chinese race 5 of the bacterial blight pathogen. Fifty-six germplasm sources were surveyed for PCR polymorphism in order to facilitate future PCR-based marker assisted transfer of bacterial blight resistance genes xa-5, xa-13 and Xa-21 to any desired varietal background which will be useful for selection of parents in breeding programmes.

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.