Targeted mutagenesis of amino acid transporter genes for rice quality improvement using the CRISPR/Cas9 system

High grain protein content(GPC) reduces rice eating and cooking quality(ECQ). We generated OsAAP6 and OsAAP10 knockout mutants in three high-yielding japonica varieties and one japonica line using the CRISPR/Cas9 system. Mutation efficiency varied with genetic background in the T_0 generation, and GPC in the T_1 generation decreased significantly,owing mainly to a reduction in glutelin content. Amylose content was down-regulated significantly in some Osaap6 and all Osaap10 mutants. The increased taste value of these mutants was supported by Rapid Visco Analysis(RVA) profiles, which showed higher peak viscosity and breakdown viscosity and lower setback viscosity than the wild type. There were no significant deficiencies in agronomic traits of the mutants. Targeted mutagenesis of OsAAP6 and OsAAP10, especially OsAAP10, using the CRISPR/Cas9 system can rapidly reduce GPC and improve ECQ of rice, providing a new strategy for the breeding cultivars with desired ECQ.

Rice Storage Proteins:Focus on Composition,Distribution,Genetic Improvement and Effects on Rice Quality

Rice storage proteins(RSPs)are plant proteins with high nutritional quality.As the second largest type of storage substance in rice,it is the main source of protein intake for people who consume rice as a staple food.The content and type of RSPs affect the appearance,processing quality and eating quality of rice.These effects involve the distribution of RSPs in rice grains as well as the interactions of RSPs with other components such as starch in rice grains.In the past two decades,some progress has been made in the genetic improvement of RSPs.However,the determination mechanism of protein content and composition in rice is still unclear,and the mechanism of the effect of RSPs on rice quality has not been elucidated.In this review,the composition,biosynthesis and distribution of RSPs,and quantitative trait loci mapping and cloning of RSP genes are summarized,the research progress of the influence of RSPs and their components on rice quality are reviewed,and the research directions in the future are proposed.

The Effect of High Temperature after Anthesis on Rice Quality and Starch Granule Structure of Endosperm

To ascertain the effect mechanism of high temperature after anthesis on rice quality, the experiment was conducted with two rice lines, the heat-tolerant line 996 and heat-sensitive line 4628, with high temperature and optimal temperature in the growth chamber to investigate the effect of high temperature stress after anthesis on rice appearance quality, milling quality, cooking and eating quality and starch granule structure of endo- sperm. The result showed that milled rice rate, head rice rate, amylose content and gel consistency of both lines decreased under high temperature stress after anthesis, while the ratio of grain length to width, chalky rate, chalkiness, protein content increased. Under high temperature treatments, gelatinization temperature, final viscosity, set back and peak time increased, breakdown decreased, Mg content and K content increased, Mg/K ra- tio decreased. Under same treatment, the extent of rice quality of heat tolerant line 996 affected by high temperature was lower than that of heat sensitive line 4628. Under high temperature stress after anthesis, starch granule arranged untightly, most of starch granules existed in the form of a single starch endosperm, refractive index decreased, transparency reduced, and led to the formation of chalk. Under high temperature stress af- ter anthesis, the increase of protein content, the decrease of Mg/K, the changes of rice RVA profile characteristics and starch granule structure of endosperm could be the main reason for the decrease of rice cooking and eating quality and appearance quality.

Effect of Interplanting with Zero Tillage and Straw Manure on Rice Growth and Rice Quality

The interplanting with zero-tillage of rice, i.e. direct sowing rice 10-20 days before wheat harvesting, and remaining about 30-cm high stubble after cutting wheat or rice with no tillage, is a new cultivation technology in wheat-rice rotation system. To study the effects of interplanting with zero tillage and straw manure on rice growth and quality, an experiment was conducted in a wheat-rotation rotation system. Four treatments, i.e. ZIS （Zero-tillage, straw manure and rice interplanting）, Zt （Zero-tillage, no straw manure and rice interplanting）, PTS （Plowing tillage, straw manure and rice transplanting）, and PT （Plowing tillage, no straw manure and rice transplanting）, were used. ZtS reduced plant height, leaf area per plant and the biomass of rice plants, but the biomass accumulation of rice at the late stage was quicker than that under conventional transplanting cultivation. In the first year （2002）, there was no significant difference in rice yield among the four treatments. However, rice yield decreased in interplanting with zero-tillage in the second year （2003）. Compared with the transplanting treatments, the number of filled grains per panicle decreased but l000-grain weight increased in interplanting with zero-tillage, which were the main factors resulting in higher yield, tnterplanting with zero-tillage improved the milling and appearance qualities of rice. The rates of milled and head rice increased while chalky rice rate and chalkiness decreased in interplanting with zero-tillage. Zero-tillage and interplanting also affected rice nutritional and cooking qualities. In 2002, ZIS showed raised protein content, decreased amylose content, softer gel consistency, resulting in improved rice quality. In 2003, zero-tillage and interplanting decreased protein content and showed similar amylose content as compared with transplanting treatments. Moreover, protein content in PTS was obviously increased in comparison with the other three treatments. The rice in interplanting with zero-tillage treatments had higher peak viscosity and breakdown, lower setback, showing better rice taste quality. The straw manure had no significant effect on rice viscosity under interplanting with zero-tillage, but had the negative influence on the rice taste quality under transplanting with plowing tillage.

Rice Quality Improvement in China

1. Demand for high quality rice in ChinaRice is the leading cereal crop that contributes about40% of the national grain production in China. The totaloutput and areas rank the first and the second position inthe world, respectively. In recent years, rice productiongrows steadily, but the quality improvement has beenlagged and the quality becomes a limiting factor. As the

Preliminary studies on rice quality of japonica-indica hybrid rice

It has been proved that high yield combina-tions can be obtained via hybridization betweenindica and japonica rice. However, people haveworried about the quality of this kind of ricefor a long time. Our researches were mainlybased on the mating pattern of "japonica malesterile line/indica restorer line" to generateand select elite japonicalinous crosses. Ricequality is shown in table 1 and 2. The scores of rice quality of 7 crossesranged from 56-59, and met the high quality

Rice quality in relation to market prices in Yunnan Province

Factors affecting rice quality and their impacts on market price were investigated in this study. On-farm survey and market survey was undertaken in three selected sites namely Kunming, Dali, and Xishuangbanna in Yunnan Province, China. Market sampling was conducted to determine important rice quality characteristics. Sixty milled rice samples were collected from domestic markets of the three sites during a period of Mar to Apr in 1994. The grain physicochemical properties of the milled rice samples were analyzed on the basis of Chinese Agricultural Ministry Standard for testing rice quality. A hedonic price model (implicit price model) was further employed to quantify relationships between quality characteristics and market prices of rice. The model can be expressed mathematically as the following function (1) This function shows that the average price paid by consumer for different grades of rice with attribute Xj. Using the ordinary least square (OLS) regression of observed market prices on measures

Effects of CO_2 enrichment and spikelet removal on rice quality under open-air field conditions

The increase of atmospheric carbon dioxide（CO_2） concentration adversely affect several quality traits of rice grains, but the biochemical mechanism remains unclear. The objectives of this study were to determine how changes in the source-sink relationship affected rice quality. Source-sink manipulation was achieved by free-air CO_2 enrichment from tillering to maturity and partial removal of spikelet at anthesis using a japonica rice cultivar Wuyunjing 23. Enrichment with CO_2 decreased the head rice percentage and protein concentration of milled rice, but increased the grain chalkiness. In contrast, spikelet removal resulted in a dramatic increase in the head rice percentage and protein concentration, and much less grain chalkiness. Neither CO_2 enrichment nor spikelet removal affected the starch content, but the distribution of starch granule size showed distinct treatment effects. O n average, spikelet removal decreased the percentage of starch granules of diameter 〉10 and 5–10 μm by 23.6 and 5.6%, respectively, and increased those with a diameter of 2–5 and 〈2 μm by 4.6 and 3.3%, respectively. In contrast, CO_2 elevation showed an opposite response： increasing the proportion of large starch granules（〉5 μm） and decreasing that of 〈5 μm. The starch pasting properties were affected by spikelet removal much more than by CO_2 elevation. These results indicated that the protein concentration and starch granule size played a role in chalkiness formation under these experimental conditions.

Effects of Milling Methods on Rice Flour Properties and Rice Product Quality:A Review

High-quality rice flour is the foundation for the production of various rice-based products.Milling is an essential step in obtaining rice flour,during which significant changes occur in the physicochemical and quality characteristics of the flour.Although rice flour obtained through mainstream wet milling methods exhibits superior quality,low production efficiency and wastewater discharge limit the development of the industry.Dry milling,on the other hand,conserves water resources,but adversely affects flour performance due to excessive heat generation.As an emerging powder-making technique,semi-dry milling offers a promising solution by enhancing flour quality and reducing environmental impact.This is achieved by minimizing soaking time through hot air treatment while reducing mechanical energy consumption to reach saturated water absorption levels.However,continuous production remains a challenge.This comprehensive review summarizes the effects of various milling technologies on rice flour properties and product qualities.It also discusses key control indicators and technical considerations for rice flour processing equipment and processes.

Effects of Different Sowing Dates and Sites on Grain Quality and RVA Profile of Nanjing 46,a Popular Cultivar of Japonica Rice(Oryza sativa L.)

[Objective] This study was performed to determine the effects of sowing dates and sites on grain quality and Rapid Viscosity Analyzer （RVA） profile of Nan-jing 46. [Method] The field experiments were carried out at five sites in Jiangsu Province, and in every site, the seeds were sowing on seven different dates. RVA related indices were measured after harvest. [Result] Along with the delay of sowing date, the mil ing quality of Nanjing 46 increased at first and then reduced, while its appearance quality went up and the cooling and edible quality decreased. With the sowing sites moving southward, the rice mil ing quality, appearance quality, and cooling and edible quality of Nanjing 46 decreased at first and then increased. Grain traits responded variably to the sowing dates and sites. The brown rice rate, grain width, grain length, length/width, mil ed rice rate and head mil ed rice rate were less affected, while the chalky rice rate and chalkiness degree were more af-fected. The gelatinization temperature, protein content, amylose content, gel consis-tency and eating value were in-between. The RVA profile analysis revealed that the breakdown value, setback value and pasting temperature were affected by the sow-ing date and geographical environment. The peak viscosity, hot viscosity, cool vis-cosity and peak time were influenced more by geographical environment. With the delay of the sowing date, peak viscosity, hot viscosity, cool viscosity, setback value and pasting temperature decreased at first and then increased, while the breakdown value changed inversely, and the peak time did not exhibit a regular trend. With the decrease of latitude, the peak viscosity, hot viscosity, breakdown and cool viscosity declined at first and then increased, the setback value and peak time decreased, while the pasting temperature increased. [Conclusion] This study wil provide refer-ences for the effective production of Nanjing 46.

Effects of nitrogen level on yield and quality of japonica soft super rice

Although studies on the balance between yield and quality of japonica soft super rice are limited, they are crucial for super rice cultivation. In order to investigate the effects of nitrogen application rate on grain yield and rice quality, two japonica soft super rice varieties, Nanjing 9108 （NJ 9108） and Nanjing 5055 （NJ 5055）, were used under seven N levels with the application rates of 0, 150, 187.5,225, 262.5, 300, and 337.5 kg ha^-1. With the increasing nitrogen application level, grain yield of both varieties first increased and then decreased. The highest yield was obtained at 300 kg ha^-1. The milling quality and protein content increased, while the appearance quality, amylose content, gel consistency, cooking/eating quality, and rice flour viscosity decreased. Milling was significantly negatively related with the eating/cooking quality whereas the appearance was significantly positively related with cooking/eating quality. These results suggest that nitrogen level significantly affects the yield and rice quality of japonica soft super rice. We conclude that the suitable nitrogen application rate for japonica soft super rice, NJ 9108 and NJ 5055, is 270 kg ha^-1, under which they obtain high yield as well as superior eating/cooking quality.

Combined effect of shading time and nitrogen level on grain filling and grain quality in japonica super rice

There is limited information about the combined effect of shading time and nitrogen （N） on grain filling and quality of rice. Therefore, two japonica super rice cultivars, Nanjing 44 and Ningjing 3, were used to study the effect of shading time and N level on the characteristics of rice panicle and grain filling as well as the corresponding yield and quality. At a low N level （150 kg N ha^-1, 150N）, grain yield decreased （by 21.07-26.07%） under the treatment of 20 days of shading before heading （BH） compared with the no shading （NS） treatment. These decreases occurred because of shortened panicle length, decreased number of primary and secondary branches, as well as the grain number and weight per panicle. At 150N, in the treatment of 20 days of shading after heading （AH）, grain yield also decreased （by 9.46-10.60%） due to the lower grain weight per panicle. The interaction of shading and N level had a significant effect on the number of primary and secondary branches. A high level of N （300 kg N ha^-1, 300N） could offset the negative effect of shading on the number of secondary branches and grain weight per panicle, and consequently increased the grain yield in both shading treatments. In superior grains, compared with 150N NS, the time to reach 99% of the grain weight （T99） was shortened by 1.6 to 1.7 days, and the grain weight was decreased by 4.18-5.91% in 150N BH. In 150N AH, the grain weight was 13.39-13.92% lower than that in 150N NS due to the slow mean and the maximum grain-filling rate （GRmean and GRmax）. In inferior grains, grain weight and GRmean had a tendency of 150N NS〉150N BH〉150N AH. Under shaded conditions, 300N decreased the grain weight due to lower GReen both in superior and inferior grains. Compared with 150N NS, the milling and appearance qualities as well as eating and cooking quality were all decreased in 150N BH and 150N AH. Shading with the high level of 300N improved the milling quality and decreased the number of chalky rice kernels, but the eating and cooking quality was reduced with increased chalky area and overall chalkiness. Therefore, in the case of short term shading, appropriate N fertilizer could be used to improve the yield and milling quality of rice, but limited application of N fertilizer is recommended to achieve good eating and cooking quality of rice.

Effects of Nitrogen Application Time on Caryopsis Development and Grain Quality of Rice Variety Yangdao 6

A pot experiment was conducted to study the effects of different nitrogen application time （during the tillering or the booting stages） with the same nitrogen rates on the caryopsis development and grain quality of rice variety Yangdao 6. The increased nitrogen fertilizer （urea）, especially applied during the booting stage, could evidently increase the milled rice rate, head rice rate and protein content in rice grains compared with the control （no nitrogen application）, and decrease chalky grain rate and amylose content. Moreover, the increased nitrogen fertilizer significantly affected the caryopsis development and enhanced the grain weight when nitrogen applied during the tillering and the booting stages, especially during the booting stage. During caryopsis development the increased nitrogen fertilizer applied during the tillering and booting stages could obviously decrease the total starch and amylose contents, but not obviously for the amylopectin content in rice grain. Increased topdressing of nitrogen fertilizer, especially applied during the booting stage, had significant effect on the development and structures of amyloplasts and proteinoplasts. That is, it could change the distribution, number and shape of amyloplasts and proteinoplasts in the endosperm cells especially in grain abdomen, Compared with the control the arrangements of amyloplasts and proteinoplasts were closer, with more numbers, higher density and less interspaces each ohter. Furthermore, most amyloplasts showed polyhedron under the increased nitrogen fertilizer level.

Effects of planting methods on yield and quality of different types of japonica rice in northern Jiangsu plain, China

Mechanical transplanting with carpet seedlings(MC) and mechanical direct seeding(MD) are newly developed planting methods, which increase in popularity and planted area each year. Knowing the difference for yield and rice quality under different planting methods is of great importance for the development of high quality and yield cultivation techniques under mechanical conditions. Therefore, three kinds of japonica rice including hybrid japonica rice, inbreed japonica rice, and soft rice were adopted as materials. And the differences in the quality of processing, appearance, cooking and eating quality, nutrition, and the rapid viscosity analyzer(RVA) profile were studied to reveal the effects of planting methods on yield and quality of different types of japonica rice. Results showed that the milled rice and head rice rates under MC was significantly higher than those under MD, and the processing quality of inbreed japonica rice was the most stable. Compared with MC, length/width ratio of rice under MD was significantly increased, and chalkiness rate, size, and degree were significantly decreased. The protein content under MD was lower than that under MC. MC showed higher peak viscosity and breakdown value than MD. The taste value was the greatest for soft rice, followed by inbreed japonica rice, and then by japonica hybrid rice, with no significant differences resulting from planting methods. Compared with MC, MD significantly improved the appearance quality, though processing quality and nutritional quality were decreased. And there was no significant difference in cooking and eating quality between MC and MD. Under different planting methods, the appearance quality of inbreed japonica rice changed the most and the processing quality was the most stable. The nutritional, cooking and eating quality of soft rice changed the least. Therefore, according to the different planting methods and market needs, selecting the appropriate rice varieties can reduce the risks in rice production and achieve good rice quality.

Difference in Grain Yield and Quality among Tillers in Rice Genotypes Differing in Tillering Capacity

A japonica variety and its iso-allelic mutant with high tillering ability were used to investigate the differences in grain yield and quality among different tillers. There was a distinct difference in panicle weight among tillers during grain filling for both genotypes, with Xiushui 11 having a greater increase rate in panicle weight, and being earlier in reaching the maximum panicle weight than the mutant. There was a great significant difference between the two genotypes in grain yield and its components, with Xiushui 11 having higher grain yield, more grains per panicle, higher filled grain percentage and grain weight than the mutant for each type of tiller. Moreover, a significant difference was found in grain yield and yield components among different tillers for both genotypes, with grain number per panicle showing the greatest variation over tillers among all yield components. Compared with Xiushui 11, the mutant had higher brown rice rate and greater ratio of length to width of brown rice, and lower chalky rice rate, amylose content, and protein content of rice. Furthermore, there was a significant difference in grain quality among tillers within a plant for both genotypes, with later initiated tillers being lower in chalky rice rate, amylose and protein contents than early initiated ones. The variation of most quality parameters among tillers within a plant was markedly larger for the mutant than for Xiushui 11.

Response of grain-filling rate and grain quality of mid-season indica rice to nitrogen application

Nitrogen is one of the important factors for high yield of rice.Apart from high yield,high quality has become the current urgent demand for rice production.Grain-filling stage is crucial for rice yield and quality formation.However,the effect of nitrogen on grain-filling characteristics and the relationship of grain-filling characteristics and rice quality of mid-season indica rice were still unclear.A field experiment was carried out to ascertain the critical grain-filling characteristics that contribute to rice milling quality,appearance quality and cooking and eating quality under nitrogen applications.The results showed that nitrogen applications prolonged the duration of superior and inferior grain filling.The mean grainfilling rate(G_(mean))and the maximum grain-filling rate(G_(max))of the inferior grains were positively correlated with chalky kernel rate,chalkiness,and amylose content.The time reaching the maximum grain-filling rate(T_(max)G)of the inferior grains was positively correlated with brown rice rate,milled rice rate,and head milled rice rate.Chalky kernel rate and chalkiness were negatively correlated with peak paste viscosity and breakdown viscosity.Less amylose content and more crude protein content were detected in nitrogen application of Liangyoupei 9 and Y Liangyou 2 both in 2016 and 2017.According to the correlation analysis,better cooking and eating quality of Y Liangyou 2 which had less amylose content might result from its higher G_(max)and G_(mean)of inferior grain than that of Liangyoupei 9 in the treatments of nitrogen application.These results indicated that the prolonging grain-filling duration and increasing grain weight at the maximum grain-filling rate of inferior grains contributed to the improvement of milling quality,appearance quality,and cooking and eating quality of mid-season indica rice under appropriate nitrogen applications.

Identification of QTLs for Cooking and Eating Quality of Rice Grain

The BIL （backcross inbred line） population derived from the cross between Koshihikari （good eating and cooking quality, japonica） and Kasalath （poor quality, indica） was used to analyze the QTLs for amylose content （AC）, gelatinization temperature （GT）, gel consistency （GC） and protein content （PC）. Eight main-effect QTLs including 2 for AC, 3 for GT, 2 for GC and 1 for PC were identified, Moreover, 27 epistatic QTL pairs including 7 for AC, 5 for GT, 4 for GC and 11 for PC were also detected while for AC and GT, one main-effect QTL with a major gene was detected, respectively. Therefore, the main-effect QTL might be more responsible for the current variation than the epistetic QTL. The result indicated that the main-effect QTL is the primary genetic basis for those traits. However, for PC, the epistatic QTL explained a much greater portion of the total variation than main-effect QTL, suggesting that epistatic loci are the primary genetic basis for such trait. In the experiment, chromosome segment substitution lines （CSSLs） were used to confirm reliabilities of the main effect QTLs detected in BIL population. Of the 8 main-effect QTLs for 4 traits in BIL analysis, 6 were confirmed end 2 remained unconfirmed by CSSLs analysis.

Effect of Nitrogen Application Rates on the Nitrogen Utilization, Yield and Quality of Rice

Research on the effect of the nitrogen application rate on the balance of the nitrogen utilization, yield and quality of rice is common in South China but is relatively lacking in Northeast China, especially in the Liaohe Delta. In this study, Yanfeng 47 rice was planted in Panjin city, China, to explore the effect of six nitrogen rates (0, 160, 210, 260, 315 and 420 kg N/ha) on the nitrogen use efficiency, rice quality and grain yield of rice plants. The results showed that the application of an appropriate nitrogen rate (210-260 kg N/ha) remarkably increased the nitrogen use efficiency of rice plants, grain yield, rice milling quality and nutritional quality and resulted in a moderate rice eating quality. Although low nitrogen rates (160 kg N/ha) maintained a high rice eating quality, they decreased grain yield and other rice qualities, and excessive nitrogen (315 kg N/ha) increased rice appearance quality but significantly reduced the nitrogen use efficiency (p < 0.05), yield and eating quality of rice. Therefore, to produce rice in the Liaohe Delta by an environmentally friendly method and guarantee rice with high quality and yield, the recommended nitrogen application rate is 210 kg N/ha.

Effect of Rolled Leaf Gene Rl_(t) on Grain Quality in Hybrid Rice

Effects of rolled leaf gene Rl（t） on grain quality characters of hybrid rice were analyzed by using three pairs of rolled leaf near-isogenic lines under two fertilizer treatments. Under normal fertilizer level （e.g. 450 kg urea per ha）, head rice rates and milled rice recovery of rolled leaf hybrids were significantly higher than those of corresponding non-rolled crosses, while the chalky rice rate and chalkiness were all lower. Of the RVA profiles, the peak viscosity, the hot paste viscosity and the breakdown viscosity of the rolled were all higher than those of the corresponding non-rolled ones to various degrees. Increasing fertilizer application for promoting panicle development increased the brown, milled and head rice rates except for Shanyou 63, furthermore, significant difference of head rice rates existed between the rolled leaf Shanyou 559 and Shanyou 559; while the peak viscosity, the hot paste viscosity and the breakdown viscosity all decreased to different levels; changes of values of other characters had no apparent regularity. It suggested that Rl（t） could improve rice quality under certain conditions.

Relationship of Parental Indica-Japonica Indexes with Yield and Grain Quality Traits of Japonica Hybrid Rice in Northern China

Taking the main parents （10 male sterile lines and 10 restorer lines） and their 100 combinations of japonica hybrid rice in northern China as materials, the relationships of parental indica-japonica indexes determined by the methods of the Cheng’s index as well as simple sequence repeat （SSR） markers with yield and grain quality traits of hybrid rice were studied. For the parents, the Cheng’s index （Chi） ranged from 13.5 to 19.3 and the indica index in SSR markers （ADi） were from 0.12 to 0.38. The classification of parents by Chi was not completely consistent with that by ADi. The Chi of male parent was more closely related to hybrid traits than that of female parent, as contrasted to ADi. At the same time, the difference between parents （PD） in Chi was more closely related to hybrid traits than that in ADi. The indica-japonica indexes of parents and their difference between parents appeared quadratic relationship to hybrid traits with the critical extremum. The directions of the correlation of indica-japonica indexes of parents and their differences with hybrid yield traits were on the opposition to those with hybrid grain quality traits. Therefore, the female parent should match the male parent moderately in indica-japonica index to obtain the optimum of hybrid traits, high yield as well as good quality.