高直链淀粉玉米amylose-extender基因功能标记的开发及应用

直链淀粉的含量是玉米淀粉品质的重要指标,受隐性核基因amylose-extender(ae)控制。生产中直链淀粉含量的测定费时、费力,因此高直链淀粉玉米的改良需要寻找快速、简便、准确的基因检测方法。本实验对高直链淀粉玉米材料和普通玉米材料ae基因的cDNA进行了测序,结果显示高直链淀粉材料cDNA序列第9外显子(84bp)全部缺失。利用该缺失设计的玉米ae基因的功能标记(ae-InDel),能够检测出纯合基因型个体(AeAe和aeae)和杂合基因型个体(Aeae),该共显性标记能有效用于高直链淀粉玉米的分子标记辅助育种。

Determination of Amylose Content and Its Relationship with RVA Profile Within Genetically Similar Cultivars of Rice(Oryza sativa L.ssp.japonica)

Anther culture pure lines with antisense Wx gene were generated by Agrobacterium tumefaciens-mediated co-transformation followed by anther culture in transgenic rice （Oryza sativa L. ssp. japonica）. The antisense Wx transgenic pure lines were used to analyze the differences and the relationship between RVA eigenvalues among the lines with different amylose contents. 77 pure lines of anther culture having antisense Wx gene were studied for the amylose content of its offspring cultivar, Wuyunjing 7. It was found that levels ofamylose content were similar to the control in 19 lines （16.0%）; 1-5% lower than control in 50 lines, of which, within 11.0-13.9% in 40 lines; and 3.1-4.0% in 8 lines. The effect of the amylose content on RVA eigenvalue was analyzed through the RVA profile graphic comparison and test of significance for RVA eigenvalues. The analysis of RVA profiles with different amylose contents indicated that there were two kinds of RVA profiles within genetically similar cultivars. The lines with 3.1-4.0% of amylose content were distinctly different from other lines and conventional glutinous rice. The comparison of the similarity curve of RVA profile of the lines with different amylose contents showed that the lines with lower amylose content had a distinguished RVA profile with the curve increasing gradually, but not exceeding the first apex. The test of significance for eigenvalues of the RVA profile in the lines with different amylose contents indicated that there were five eigenvalues remarkably different among the lines with 3.1-4.0% of amylose content compared with other two groups. Further, there were three eigenvalues remarkably different among the lines with 11.0-13.5% of amylose content compared with control group. An optimized mathematical model, which characterizes the relationships between the amylose content and RVA eigenvalue, has been established in this study. From this model, it was clearly indicated that the parameters PeT and SBV were more related to amylose content. It was concluded that the differences in amylose content not only affected the eigenvalues of RVA but also caused different RVA profiles within genetically similar cultivars. The introduction of antisense Wx gene could lead to reduce the amylose content and lay theoretical foundation for quality improvement in rice breeding programs.

Expression Profiles and Protein Complexes of Starch Biosynthetic Enzymes from White-Core and Waxy Mutants Induced from High Amylose Indica Rice

Physicochemical properties of endosperm starches in milled rice determine its cooking and eating quality.Amylose is synthesized by granule-bound starch synthase I(GBSSI),whilst amylopectin is synthesized by the synergistic activities of starch synthases(SSs),branching enzymes(BEs)and debranching enzymes(DBEs).However,the complexes formed by starch biosynthetic enzymes are not well characterized.Gene expression profiles and protein complexes were determined in white-core(GM645)and waxy(GM077)mutants derived from a high amylose indica rice Guangluai 4(GLA4).In GM645,genes including AGPS1,GBSSI,SSIIa,BEI,BEIIa,BEIIb,PUL,ISA1 and SP were significantly downregulated during seed development.In GM077,the expression levels of AGPL2,AGPS1,AGPS2b,SSIIIa,BEI,PUL and ISA1 were significantly upregulated.Co-immunoprecipitation assays revealed interactions of SSs-BEs,SSs-PUL and BEs-PUL in developing seeds.However,weak SSI-SSIIa interaction was detected in GM077,whilst SSI-PUL interaction was absent.Weak interaction signals for SSI-SSIIa,SSIIa-BEI,SSIIa-BEIIb,BEI-BEIIb and SSI-BEI were also observed in GM645.These results suggest that the protein-protein interactions for starch biosynthesis are modified in mutants,which provides insight into the mechanisms of starch biosynthesis,particularly in indica rice.

Application of Near Infrared Transmittance Spectroscopy(NITS)in Determining Amylose Content of Rice Grain

Rice samples were scanned with a Foss Tecator Infratec and their amylose content was determinedusing a standard wet chemical method.Calibration software along with several mathematicaltreatments of the sample spectra and several regression methods was used to optimize a nearinfrared transmittance spectroscopy equation for determining rice amylose content. The standarderror of calibration, standard error of cross-validation and regression squared of thecalibration were 1.489, 1.761 and 0.885,respectively.

Progresses in Researches of the Application of Low-Amylose Content Rice Gene for Breeding

Amylose content is a key determinant of eating quality of rice. With the characteristics offluffy texture, glossy appearance when cooked, remaining soft when cooled and excellentpuffing ability, the low-amylose rice with amylose content 5-15% could be served as not onlycooked rice directly, but also good material for convenience, mixed rice and puffing foods.Current status on characterization, inheritance, molecular mechanism and breeding of low-amylose content rice was reviewed in this paper, strategy of related researches in the era ofglymics was mainly discussed furthermore. The future research should focus on screening andenhancing the germplasm, further elucidating the molecular mechanism on mutation of lowamylose content, utilizing the genes independent of Wx on low-amylose content rice breedingprogram, and developing high quality functional rice cultivars for special usage throughpyramiding low amylose gene and other special quality genes.

Comparison of Near Infrared Spectroscopy Models for Determining Protein and Amylose Contents Between Calibration Samples of Recombinant Inbred Lines and Conventional Varieties of Rice

The near infrared spectra of 178 recombinant inbred lines （RILs） from the cross of Ⅱ-32B/Yuezaoxian 6 （YZX6） and 511 varieties in rice were acquired. A total of 80 RILs and 96 cultivars were selected as modeling samples by comparing the spectra similarity primarily. Three partial least square （PLS） regression models were developed, based on the RILs （RIL-model）, the varieties （Var-model） and their mixture （Mix-model）, for protein content （PC） and amylose content （AC）, respectively. Cross validation and outer prediction showed that the models were largely influenced by the range and distribution of modeling samples. The regression model of PC based on the cultivars and the model of AC based on RILs had higher coefficient of determination （r^2 ≥ 0.9） and lower root mean square error of cross validation （RMSECVs）. The disadvantages of RIL samples for PC model and variety samples for AC model were probably caused by the narrow range of variance. Aberrant predictions were obtained for outer sample with PC or AC outside the range or within the distribution gap of modeling samples. The Mix-models gave more reliable prediction as the distribution of RIL and variety modeling samples were complementary to each other.

Relationship Between the First Base of the Donor Splice Site of Waxy Gene Intron 1 and Amylose Content in Yunnan Indigenous Rice Varieties

There exists a single nucleotide polymorphism, G or T, at the first base of the donor splice site of waxy gene intron 1 in rice. In order to study the relationship between the first base of the donor splice site of waxy gene intron 1 and amylose content in rice, the one-step PCR method was used to determine whether it is G or T in 220 Yunnan indigenous rice varieties from 14 districts, 55 towns/counties of Yunnan Province, and 101 varieties of which were validated by the PCR-Acc I method. According to the G/T polymorphism, 164 rice varieties showed GG-genotype, while the other 56 fell into TT- genotype, accounting for 74.5% and 25.5% of all the test varieties, respectively. When all the rice varieties were divided into indica and japonica subspecies, it was found that 80.5% of indica rice and 67.0% of japonica rice belonged to GG-genotype. The rice varieties with GG-genotype had significantly higher amylose content （18.95% on average） than those with TT- genotype （all below 16%）, but 33 rice varieties with GG-genotype still had low amylose content ranging from 3.91% to 15.93%, and most of them came from the Dai minority area in the Southwest of Yunnan Province. However, there was no significant difference in the mean amylose content of the same GG or TT genotypes between indica and japonica rice, suggesting that different genetic backgrounds, indica or japonica, had no effect on amylose content. The coefficient of correlation between the genotype and amylose content was 0.733 （P〈0.01）.

Effects of dietary amylose to amylopectin ratio on growth performance,carcass quality characteristics and meat fatty acids in Chinese Qinchuan cattle

This study evaluated the effects of the dietary starch amylose/amylopectin ratio on growth performance,rumen fermentation and blood parameters,carcass characteristics and marbling score,and meat fatty acid profiles and transcriptional changes in the genes involved in the gluconeogenesis pathway in Chinese Qinchuan cattle.Forty-five cattle were randomly divided into three groups.The bulls were fed a control diet(middle amylose/amylopectin ratio=0.47)or diets with either waxy corn starch(WS)inclusion(low amylose/amylopectin ratio=0.23)or pea starch(PS)inclusion(high amylose/amylopectin ratio=0.60)for 90 days.The bulls were individually allowed to receive the diets at the daily amount of at least 2.1%of their individual BW twice daily.The bulls were weighed at 45-day intervals,and blood and rumen fluid samples were also collected at 45-day intervals.The bulls were slaughtered and longissimus thoracis(LT)samples were collected for meat quality measurements.Compared with PS,the average daily gain(ADG)and feed efficiency in control and WS were increased(P<0.05)during d 0 to 45.However,the feed efficiency in PS was increased(P<0.05)compared with the control and WS during d 46 to 90.During the whole feeding trial,the ADG in control and WS showed trends that were higher(0.05<P<0.1)than that of PS,while other growth performance measures and feed intake did not differ(P>0.05)among the three groups.Compared with PS,the serum insulin concentration in WS was increased(P<0.05)at d 45,but not at d 90.Compared with control and WS,the serum SOD and T-AOC concentrations in PS were increased(P<0.05)at d 45,but not at d 90.Compared with control,the rumen microbial crude protein contents in WS and PS were increased(P<0.05)at d 0,while those of control and WS were higher(P<0.05)than that of PS at d 45,but not at d 90.Compared with PS,the backfat thickness in control and WS were increased(P<0.05),however,the marbling score and expression of genes related to the gluconeogenesis pathway in liver of the three groups did not differ(P>0.05).Chemical and physio-chemical compositions of LT did not differ(P>0.05)among the three groups.In conclusion,diets with low or middle amylose/amylopectin ratios increased the backfat thickness,and tended to improve ADG,but had no effect on other carcass characteristics,fat content in the LT,or the expression of hepatic genes related to the gluconeogenesis pathway in Chinese Qinchuan cattle.

The effect of amylose on kernel phenotypic characteristics,starch-related gene expression and amylose inheritance in naturally mutated high-amylose maize

High-amylose maize starch has great pote ntial for widespread in dustrial use due to its ability to form strong gels and film and in the food processi ng field,thus servi ng as a resista nt starch source.However,there is still a substa ntial shortage of high-amylose maize due to the limitation of natural germplasm resources,although the well-known amylose extender(ae)gene mutants have bee n found to produce high-amylose maize lines since 1948.In this con text,high-amylose maize lines(13 inbreds and 18 hybrids)originating from a natural amylose mutant in our testing field were utilized to study the correlation between amylose content(AC)and phenotypic traits(kernel morphology and endosperm glossiness),grain filling characteristics,gene expression,and amylose inheritanee.Our results showed that AC was negatively correlated with total starch con tent but was not correlated with grain phe no types,such as kernel full ness,kernel morphology and endosperm glossiness.Maize lines with higher amylose had a greater grain filling rate than that of the controI(B73)during the first 20 days after pollination(DAP).Both starch debranching enzyme(DBE)groups and starch branching enzyme lib(SBEIIb)groups showed a greater abundance in the control(B73)than in the high-amylose maize lines.Male parents directly predicted AC of Fv which was moderately positively correlated with the F2 generation.

Insights into the swelling process and drug release mechanisms from cross-linked pectin/high amylose starch matrices

Cross-linked pectin/high amylose mixtures were evaluated as a new excipient for matrix tablets formulations,since the mixing of polymers and cross-linking reaction represent rational tools to reach materials with modulated and specific properties that meet specific therapeutic needs.Objective:In this work the influence of polymer ratio and cross-linking process on the swelling and the mechanism driving the drug release from swellable matrix tablets prepared with this excipient was investigated.Methods:Cross-linked samples were characterized by their micromeritic properties(size and shape,density,angle of repose and flow rate)and liquid uptake ability.Matrix tablets were evaluated according their physical properties and the drug release rates and mechanisms were also investigated.Results:Cross-linked samples demonstrated size homogeneity and irregular shape,with liquid uptake ability insensible to pH.Cross-linking process of samples allowed the control of drug release rates and the drug release mechanism was influenced by both polymer ratio and cross-linking process.The drug release of samples with minor proportion of pectin was driven by an anomalous transport and the increase of the pectin proportion contributed to the erosion of the matrix.Conclusion:The cross-linked mixtures of high amylose and pectin showed a suitable excipient for slowing the drug release rates.

Amylose Content,Morphology,Crystal Structure,and Thermal Properties of Starch Grains in Main and Ratoon Rice Crops

Rice ratooning,or the production of a second rice crop from stubble after the harvest of the main crop,is considered to be a green and resource-efficient rice production system.The present study was conducted to examine variance in amylose content(AC),grain morphology,crystal structure,and thermal properties of starch between main-and ratoon-season rice of seven varieties.Ratoon-season rice grains had higher ACs and significantly lower transition gelatinization temperatures(To,Tp,and Tc)than did main-season rice grains.The relative crystallinity and lamellar peak intensity of ratoon-season rice starch were 7.89%and 20.38%lower,respectively,than those of main-season rice starch.In addition,smaller granules with smoother surfaces and lower thermal parameters were observed in the starch of ratoon-season rice.The relative crystallinity and lamellar peak intensity of starch correlated negatively with the AC and positively with transition gelatinization temperatures.These results suggest that the superior cooking quality of ratoon-season rice is attributable to the moderate increase of grain AC,which reduces the relative crystallinity,weakens the crystal structure,and lead to a decrease in the gelatinization temperature.

Molecular marker for screening rice cultivars with intermediate amylose content in indica

The waxy(Wx)gene of rice encodes the granule-bound starch synthase(GBSS),which was required in the synthesis of amylose in endosperm.Theamylose content(AC)of rice endosperm played an important role in grain yield,palatability,and processing quality.Our previous researehes showed that en-dosperm AC and GBSS contents were correlated with the ability of excising intron1 from the leader sequence of the Wx transcript.Cultivars with high endosperm

Lowering Grain Amylose Content in Backcross Offsprings of indica Rice Variety 057 by Molecular Marker-Assisted Selection

To lower the amylose content （AC） of the indica rice restorer line 057 with high AC, backcrosses were made respectively by using four indica varieties （R367, 91499, Yanhui 559, Hui 527） as low AC donor parents and 057 as the recurrent parent. A molecular marker （PCR-Acc Ⅰ） was used to identify the genotypes （GG, TT and GT） of the waxy （Wx） gene. Plants with GT genotype were selected and used as female parent and crossed with 057 to advance generation. The ACs of rice grains harvested from plants with different Wx genotypes were measured and compared to analyze the efficiency of marker-assisted selection. The ACs of the rice grain, harvested from the plants of Wx genotypes GG, GT and TT, were higher than 20%, in the range of 17.7-28.5%, and less than 18%, respectively. The PCR-Acc Ⅰ marker could be used for efficiently lowering the AC of 057 through backcrossing, and there were some influence of parental genetic background on the AC of rice grains with the same Wx genotype.

Amylose Content and Starch Granule Size in Rice Grains are Affected By Growing Season

Amylose content and starch granule size in grains influence rice quality,which differs between the early(ES)and late season(LS).The objective of this study was to determine the variation of amylose content and starch granule size between seasons and find the main reasons(e.g.,temperature and solar radiation)for the observed variation.Field experiments with six rice varieties(three high and three low amylose content rice)planted in the ES and LS were conducted in 2016 and 2017,respectively.The mean temperatures during the filling stage were higher in ES,however,the daily temperatures at 7-10 days after flowering(DAF)in 2016,and at 5-10,13-14 DAF in 2017 were higher in LS.The results showed that amylose content in LS was lower than in ES with high amylose content rice varieties(HACV);the opposite trend occurred with low amylose content rice varieties(LACV).The mean starch granule diameter was higher in LS than ES in 2016,but the opposite result occurred in 2017 with all rice.Our results suggest that higher temperatures increased and decreased the amylose content in HACV and LACV,respectively.Temperatures at 5-15 DAF were important for the formation of starch granules:lower temperatures during 10-14 DAF increased the proportion of larger starch granules(d>6.21μm,some with d>13.3μm),and higher temperatures at 5-6 DAF increased the proportion of starch granules with diameter 4.24-6.21μm.

Hydrogen Bonds Sites of Amylose or Amylopectin from Starch at the ONION Level (B3LYP/6-311++G [d, p]: AM1)

The research aims to help reduce the enormous post-harvest losses of rice or bananas in Cote d’Ivoire. It focuses on the breakdown mechanism of their starch. This process contributes to their decay. This study wants to elucidate it;to do this, it analyzes the tetra or penta saccharide reactions with the water or dioxide carbon. It calculates the geometric, energetic and spectroscopic parameters at the ONIOM level (B3LYP/6-311++G [d, p]: AM1). These quantities allow establishing that oxygen <img src="Edit_7761c26d-f67d-4826-ab0e-8bff55d6faa3.png" alt="" /> represents the privileged hydrogen bond (HB) site for amylose with four or five D-glucose building blocks. They suggest <img src="Edit_ecd55400-653e-43d0-986d-9814259ebda7.png" alt="" /> and <img src="Edit_048b683f-5690-4078-b8ad-88fa25c5875c.png" alt="" /> are respectively those of amylopectin when the latter consists of four and five D-glucose synthons. They prove that amylose deteriorates before it;the degradation of starch begins with its alteration into disaccharides. It continues with that of amylopectin into tri or tetra-saccharides with four or five building blocks.

A New and Enantioselective Chromatographic Method for Linagliptin on Amylose Coated Chiral Stationary Phase

A new and enantioselective liquid chromatographic method was developed for estimation of S-Linagliptin in Linagliptin (LINA) drug substances. The desired enantiomeric separation was achieved on Chiralpak AD-H (250*4.6 mm*5 μm) column with the mobile phase composition of ethanol, methanol and diethylamine in a ratio of 90:10:0.1 (v/v/v) with flow rate of 0.5 mL·min-1 and column oven temperature 30°C and the eluted compounds were monitored at 225 nm. In the proposed chiral method, USP resolutions between both the enantiomers were more than 5.0. Limit of detection and Limit of quantitation of S-LINA was found to be 0.03 μg·mL-1 and 0.10 μg·mL-1 respectively. Linearity study was conducted from LOQ to 150% and correlation coefficient found to be 0.9997. Accuracy was within the range of 98.6% to 101.5%. To prove selectivity power of the method specificity study was conducted by subjecting drug substance to acid, base, hydrolysis, oxidation and photolysis and ensured the peak purity of analyte in degraded samples. Moreover, the method has been fully validated as per ICH guidelines. The proposed method is precise, accurate, linear, rugged, robust and suitable for accurate quantification of S-LINA in LINA drug substance.

Analysis of genetic effects and heritabilities for amylose content of indica rice in different environments

An understanding of the genetic effects of em-bryo genes for rice quality traits is of impor-tance for rice quality improvement. Analysisof embryo, endosperm, cytoplasmic, and ma-ternal genetic effects and genotype×environ-

基于无人机多光谱影像的寒地水稻品质估测

【目的】在稻谷收获前准确估测稻米品质,为改善水稻养分管理、实现优质优价提供参考。【方法】试验在黑龙江省佳木斯市大兴农场和青龙山农场进行,两个地点土壤肥力具有较大差异。每个农场选择一块稻田,以20 m×20 m间隔采集土壤样品分析碱解氮含量,通过变量施氮形成产量水平、长势差异明显的水稻群体。利用大疆精灵4四旋翼多光谱无人机获取水稻分蘖期、拔节期、抽穗期和成熟期冠层多光谱数据。成熟期采集长势具有明显差异的水稻样品和对应的土壤样品,测定土壤有机质和速效氮、磷、钾含量,水稻籽粒蛋白质和直链淀粉含量,产量和食味值,随机选择其中67%的数据通过逐步多元线性回归构建不同生育时期的品质估算模型,其余33%的数据用于模型的验证。【结果】大兴农场水稻成熟期土壤有机质、碱解氮、有效磷和速效钾含量的变异系数分别为11.65%、14.44%、37.66%和11.60%,青龙山农场分别为14.45%、14.32%、36.37%和28.51%。成熟期两个地点水稻产量和食味值的变异系数均大于10%,青龙山农场水稻蛋白质含量变异系数也超过10%,而两地稻米直链淀粉含量的变异系数仅为1.11%~1.83%,因此,水稻直链淀粉含量不适合用于后续的品质估测分析。分蘖期至抽穗期,大兴农场和青龙山农场水稻蛋白质多元回归模型的R~2和RMSE分别为0.262~0.794和0.259~0.686,食味值多元回归模型的R~2和RMSE分别为0.240~0.755和4.211~7.588,估测效果不理想。成熟期,大兴农场和青龙山农场水稻蛋白质多元回归模型的R~2和RMSE分别为0.791~0.874和0.166~0.365,食味值多元回归模型的R~2和RMSE分别为0.786~0.852和2.836~4.039。成熟期基于植被指数的蛋白质和食味值估测精确度优于抽穗期。在抽穗期,将土壤pH、有机质、碱解氮、有效磷、速效钾等指标与植被指数一起建立与蛋白质和食味值的多元回归模型,大兴农场水稻蛋白质模型的R~2由0.585提高到0.720,RMSE由0.301降低至0.247;两农场食味值模型的R~2由0.565~0.755提高到0.706~0.787,RMSE由4.318~4.854降低至3.993~4.029,基本可以满足对稻米品质估测的精度,而在成熟期加入上述土壤指标提升精度不明显。【结论】水稻成熟期利用多光谱无人机对稻米品质进行原位估测的精度较高,如果将估测时期提前到抽穗期,除植被指数外,还需配合土壤有机质和速效养分含量数据来提高多光谱无人机预测稻米品质的精度。