Creating a zero amylose barley with high soluble sugar content by genome editing.

Amylose biosynthesis is strictly associated with granule-bound starch synthase I (GBSSI) encoded by the Waxy gene. Mutagenesis of single bases in the Waxy gene, which induced by CRISPR/Cas9 genome editing, caused absence of intact GBSSI protein in grain of the edited line. The amylose and amylopectin contents of waxy mutants were zero and 31.73%, while those in the wild type were 33.50% and 39.00%, respectively. The absence of GBSSI protein led to increase in soluble sugar content to 37.30% compared with only 10.0% in the wild type. Sucrose and ß-glucan, were 39.16% and 35.40% higher in waxy mutants than in the wild type, respectively. Transcriptome analysis identified differences between the wild type and waxy mutants that could partly explain the reduction in amylose and amylopectin contents and the increase in soluble sugar, sucrose and ß-glucan contents. This waxy flour, which showed lower final viscosity and setback, and higher breakdown, could provide more option for food processing.

Dynamics of starch formation and gene expression during grain filling and its possible influence on grain quality.

In rice, grain filling is a crucial stage where asynchronous filling of the pollinated spikelet's of the panicle occurs. It can influence both grain quality and yield. In rice grain, starch is the dominant component and contains amylose and amylopectin. Amylose content is the chief cooking quality parameter, however, rice varieties having similar amylose content varied in other parameters. Hence, in this study, a set of varieties varying in yield (04) and another set (12) of varieties that are similar in amylose content with variation in gel consistency and alkali spreading value were used. Panicles were collected at various intervals and analysed for individual grain weight and quantities of amylose and amylopectin. Gas exchange parameters were measured in varieties varying in yield. Upper branches of the panicles were collected from rice varieties having similar amylose content and were subjected to gene expression analysis with fourteen gene specific primers of starch synthesis. Results indicate that grain filling was initiated simultaneously in multiple branches. Amylose and amylopectin quantities increased with the increase in individual grain weight. However, the pattern of regression lines of amylose and amylopectin percentages with increase in individual grain weight varied among the varieties. Gas exchange parameters like photosynthetic rate, stomatal conductance, intercellular CO2 and transpiration rate decreased with the increase in grain filling period in both good and poor yielding varieties. However, they decreased more in poor yielders. Expression of fourteen genes varied among the varieties and absence of SBE2b can be responsible for medium or soft gel consistency.

The Ruminococcus bromii amylosome protein Sas6 binds single and double helical α-glucan structures in starch.

Resistant starch is a prebiotic accessed by gut bacteria with specialized amylases and starch-binding proteins. The human gut symbiont Ruminococcus bromii expresses Sas6 (Starch Adherence System member 6), which consists of two starch-specific carbohydrate-binding modules from family 26 (RbCBM26) and family 74 (RbCBM74). Here, we present the crystal structures of Sas6 and of RbCBM74 bound with a double helical dimer of maltodecaose. The RbCBM74 starch-binding groove complements the double helical α-glucan geometry of amylopectin, suggesting that this module selects this feature in starch granules. Isothermal titration calorimetry and native mass spectrometry demonstrate that RbCBM74 recognizes longer single and double helical α-glucans, while RbCBM26 binds short maltooligosaccharides. Bioinformatic analysis supports the conservation of the amylopectin-targeting platform in CBM74s from resistant-starch degrading bacteria. Our results suggest that RbCBM74 and RbCBM26 within Sas6 recognize discrete aspects of the starch granule, providing molecular insight into how this structure is accommodated by gut bacteria.

Effect of non-starch components on the structural properties, physicochemical properties and in vitro digestibility of waxy highland barley starch.

Highland barley (HB) endosperm with an amylose content of 0-10 % is called waxy HB (WHB). WHB is a naturally slow-digesting grain, and the interaction between its endogenous non-starch composition and the WHB starch (WHBS) has an important effect on starch digestion. This paper focuses on the mechanisms by which the components of ß-glucan, proteins and lipids affect the molecular, granular, crystalline structure and digestive properties of WHBS. After eliminating the main nutrients except for starch, the estimated glycemic index (eGI) of the samples rose from 62.56 % to 92.93 %, and the rapidly digested starch content increased from 60.81 % to 98.56 %, respectively. The resistant starch (RS) content, in contrast, dropped from 38.61 % to 0.13 %. Comparatively to lipids, ß-glucan and protein contributed more to the rise in eGI and decline in RS content. The crystalline characteristics of starch were enhanced in the decomposed samples. The samples' gelatinization properties improved, as did the order of the starch molecules. Protein and ß-glucan form a dense matrix on the surface of WHBS particles to inhibit WHBS digestion. In summary, this study revealed the mechanism influencing the digestibility of WHBS from the perspective of endogenous non-starch composition and provided a theoretical basis to develop slow-digesting foods.

Mechanism of action of three different glycogen branching enzymes and their effect on bread quality.

Bread staling adversely affects the quality of bread, but starch modification by enzymes can counteract this phenomenon. Glycogen branching enzymes (GBEs) used in this study were isolated from Deinococcus geothermalis (DgGBE), Escherichia coli (EcGBE), and Vibrio vulnificus (VvGBE). These enzymes were characterized and applied for starch dough modification to determine their role in improving bread quality. First, the branching patterns, activity on amylose and amylopectin, and thermostability of the GBEs were determined and compared. EcGBE and DgGBE exhibited better thermostable characteristics than VvGBE, and all GBEs exhibited preferential catalysis of amylopectin over amylose but different degrees. VvGBE and DgGBE produced a large number of short branches. Three GBEs degraded the starch granules and generated soluble polysaccharides. Moreover, the maltose was increased in the starch slurry but most significantly in the DgGBE treatment. Degradation of the starch granules by GBEs enhanced the maltose generation of internal amylases. When used in the bread-making process, DgGBE and VvGBE increased the dough and bread volume by 9 % and 17 %, respectively. The crumb firmness and retrogradation of the bread were decreased and delayed significantly more in the DgGBE bread. Consequently, this study can contribute to understanding the detailed roles of GBEs in the baking process.

Evaluation of altered starch mutants and identification of candidate genes responsible for starch variation in wheat.

BACKGROUND: Induction of mutation through chemical mutagenesis is a novel approach for preparing diverse germplasm. Introduction of functional alleles in the starch biosynthetic genes help in the improvement of the quality and yield of cereals. RESULTS: In the present study, a set of 350 stable mutant lines were used to evaluate dynamic variation of the total starch contents. A megazyme kits were used for measuring the total starch content, resistant starch, amylose, and amylopectin content. Analysis of variance showed significant variation (p < 0.05) in starch content within the population. Furthermore, two high starch mutants (JE0173 and JE0218) and two low starch mutants (JE0089 and JE0418) were selected for studying different traits. A multiple comparison test showed that significant variation in all physiological and morphological traits, with respect to the parent variety (J411) in 2019-2020 and 2020-2021. The quantitative expression of starch metabolic genes revealed that eleven genes of JE0173 and twelve genes of JE0218 had consistent expression in high starch mutant lines. Similarly, in low starch mutant lines, eleven genes of JE0089 and thirteen genes of JE0418 had consistent expression in all stages of seed development. An additional two candidate genes showed over-expression (PHO1, PUL) in the high starch mutant lines, indicating that other starch metabolic genes may also contribute to the starch biosynthesis. The overexpression of SSII, SSIII and SBEI in JE0173 may be due to presence of missense mutations in these genes and SSI also showed overexpression which may be due to 3-primer_UTR variant. These mutations can affect the other starch related genes and help to increase the starch content in this mutant line (JE0173). CONCLUSIONS: This study screened a large scale of mutant population and identified mutants, could provide useful genetic resources for the study of starch biosynthesis and genetic improvement of wheat in the future. Further study will help to understand new genes which are responsible for the fluctuation of total starch.

The OsNAC24-OsNAP protein complex activates OsGBSSI and OsSBEI expression to fine-tune starch biosynthesis in rice endosperm.

Starch accounts for up to 90% of the dry weight of rice endosperm and is a key determinant of grain quality. Although starch biosynthesis enzymes have been comprehensively studied, transcriptional regulation of starch-synthesis enzyme-coding genes (SECGs) is largely unknown. In this study, we explored the role of a NAC transcription factor, OsNAC24, in regulating starch biosynthesis in rice. OsNAC24 is highly expressed in developing endosperm. The endosperm of osnac24 mutants is normal in appearance as is starch granule morphology, while total starch content, amylose content, chain length distribution of amylopectin and the physicochemical properties of the starch are changed. In addition, the expression of several SECGs was altered in osnac24 mutant plants. OsNAC24 is a transcriptional activator that targets the promoters of six SECGs; OsGBSSI, OsSBEI, OsAGPS2, OsSSI, OsSSIIIa and OsSSIVb. Since both the mRNA and protein abundances of OsGBSSI and OsSBEI were decreased in the mutants, OsNAC24 functions to regulate starch synthesis mainly through OsGBSSI and OsSBEI. Furthermore, OsNAC24 binds to the newly identified motifs TTGACAA, AGAAGA and ACAAGA as well as the core NAC-binding motif CACG. Another NAC family member, OsNAP, interacts with OsNAC24 and coactivates target gene expression. Loss-of-function of OsNAP led to altered expression in all tested SECGs and reduced the starch content. These results demonstrate that the OsNAC24-OsNAP complex plays key roles in fine-tuning starch synthesis in rice endosperm and further suggest that manipulating the OsNAC24-OsNAP complex regulatory network could be a potential strategy for breeding rice cultivars with improved cooking and eating quality.

Distribution of Aromatic Amino Acid Residues in Substrate-Binding Regions Modulates Substrate Specificity of Microbial Debranching Enzymes.

Debranching enzymes (DBEs) directly hydrolyze α-1,6-glucosidic linkages in glycogen, starch, and related polysaccharides, making them important in the starch processing industry. However, the ambiguous substrate specificity usually restricts synergistic catalysis with other amylases for improving starch utilization. Herein, a glycogen-debranching enzyme from Saccharolobus solfataricus (SsGDE) and two isoamylases from Pseudomonas amyloderamosa (PaISO) and Chlamydomonas reinhardtii (CrISO) were used to investigate the molecular mechanism of substrate specificity. Along with the structure-based computational analysis, the aromatic residues in the substrate-binding region of DBEs played an important role in binding substrates. The aromatic residues in SsGDE appeared clustered, contributing to a small substrate-binding region. In contrast, the aromatic residues in isoamylase were distributed dispersedly, forming a large active site. The distinct characteristics of substrate-binding regions in SsGDE and isoamylase might explain their substrate preferences for maltodextrin and amylopectin, respectively. By modulating the substrate-binding region of SsGDE, variants Y323F and V375F were obtained with significantly enhanced activities, and the activities of Y323F and V375F increased by 30 and 60% for amylopectin, and 20 and 23% for DE4 maltodextrin, respectively. This study revealed the molecular mechanisms underlying the substrate specificity for SsGDE and isoamylases, providing a route for engineering enzymes to achieve higher catalytic performance.

Authentication of ST25 rice using temperature-perturbed Raman measurement with variable selection by Incremental Association Markov Blanket.

A temperature-perturbed transmission Raman measurement was demonstrated for the discrimination of ST25 and non-ST25 rice samples. ST25 rice is a premium long-grain Vietnamese rice with the aroma of pandan leaves and the scent of early sticky rice. Raman spectra of rice samples were acquired with temperature perturbation ranging from 20 to 50 °C, and the variables (intensities of peaks) with greater discrimination were selected from the spectra using Incremental Association Markov Blanket (IAMB) for authentication. The combination of four, seven, and four variables selected from the spectra at 20, 30, and 50 °C, respectively, yielded the highest accuracy of 97.9%. The accuracies in the single-temperature measurements were lower, suggesting that the combination of mutually complementary spectral features acquired at these temperatures is synergetic to recognize the compositional differences between two sample groups, such as in the amylose/amylopectin ratio and the protein constituent.

LIKE EARLY STARVATION 1 and EARLY STARVATION 1 promote and stabilize amylopectin phase transition in starch biosynthesis.

Starch, the most abundant carbohydrate reserve in plants, primarily consists of the branched glucan amylopectin, which forms semi-crystalline granules. Phase transition from a soluble to an insoluble form depends on amylopectin architecture, requiring a compatible distribution of glucan chain lengths and a branch-point distribution. Here, we show that two starch-bound proteins, LIKE EARLY STARVATION 1 (LESV) and EARLY STARVATION 1 (ESV1), which have unusual carbohydrate-binding surfaces, promote the phase transition of amylopectin-like glucans, both in a heterologous yeast system expressing the starch biosynthetic machinery and in Arabidopsis plants. We propose a model wherein LESV serves as a nucleating role, with its carbohydrate-binding surfaces helping align glucan double helices to promote their phase transition into semi-crystalline lamellae, which are then stabilized by ESV1. Because both proteins are widely conserved, we suggest that protein-facilitated glucan crystallization may be a general and previously unrecognized feature of starch biosynthesis.

Evaluation of pea/rice and amylopectin/chromium as an alternative protein source to improve muscle protein synthesis in rats.

BACKGROUND: A preclinical study reported that the combination of an amylopectin/chromium complex (ACr) of branched-chain amino acids (BCAA) significantly enhanced muscle protein synthesis (MPS). This study was conducted to determine the effects of the addition of ACr complex to a pea/rice (PR) protein on MPS, insulin, muslin levels, and the mTOR pathway in exercised rats. METHODS: Twenty-four rats were divided into three groups: (i) exercise (Ex); (ii) Ex + PR 1:1 blend (0.465 g/kg BW); (iii) Ex + PR + ACr (0.155 g/kg BW). On the day of single-dose administration, after the animals were exercised at 26/m/min for 2 h, the supplement was given by oral gavage. The rats were injected with a bolus dose (250 mg/kg BW, 25 g/L) of deuterium-labeled phenylalanine to determine the protein fractional synthesis rate (FSR) one h after consuming the study product. RESULTS: The combination of PR and ACr enhanced MPS by 42.55% compared to the Ex group, while Ex + PR alone increased MPS by 30.2% over the Ex group (p < 0.0001) in exercised rats. Ex + PR plus ACr significantly enhanced phosphorylation of mTOR and S6K1 (p < 0.0001), and 4E-BP1 (p < 0.001) compared to the Ex (p < 0.0001). PR to ACr also significantly increased insulin and musclin levels (p < 0.0001) in exercised rats. Additionally, compared to Ex + PR alone, Ex + PR + ACr enhanced mTOR (p < 0.0001) and S6K1 (p < 0.0001) levels. CONCLUSION: These data suggested that PR + ACr may provide an alternative to animal proteins for remodeling and repairing muscle by stimulating MPS and mTOR signaling pathways in post-exercised rats. More preclinical and clinical human studies on combining pea/rice and amylopectin/chromium complex are required.

Genetic Engineering of Starch Biosynthesis in Maize Seeds for Efficient Enzymatic Digestion of Starch during Bioethanol Production.

Maize accumulates large amounts of starch in seeds which have been used as food for human and animals. Maize starch is an importantly industrial raw material for bioethanol production. One critical step in bioethanol production is degrading starch to oligosaccharides and glucose by α-amylase and glucoamylase. This step usually requires high temperature and additional equipment, leading to an increased production cost. Currently, there remains a lack of specially designed maize cultivars with optimized starch (amylose and amylopectin) compositions for bioethanol production. We discussed the features of starch granules suitable for efficient enzymatic digestion. Thus far, great advances have been made in molecular characterization of the key proteins involved in starch metabolism in maize seeds. The review explores how these proteins affect starch metabolism pathway, especially in controlling the composition, size and features of starch. We highlight the roles of key enzymes in controlling amylose/amylopectin ratio and granules architecture. Based on current technological process of bioethanol production using maize starch, we propose that several key enzymes can be modified in abundance or activities via genetic engineering to synthesize easily degraded starch granules in maize seeds. The review provides a clue for developing special maize cultivars as raw material in the bioethanol industry.

Suitable nitrogen fertilizer application drives the endosperm development and starch synthesis to improve the physicochemical properties of common buckwheat grain.

The effects of nitrogen (N) fertilizer on endosperm development, starch component, key enzyme activity and grain quality of common buckwheat were investigated in this study. The results showed that N fertilization significantly enhanced the number and area of endosperm cells, and significant increases were also observed in the contents of amylose, amylopectin and total starch. With increasing N level, the activities of key enzyme significantly increased showing the maximum under the N2 level (180 kg N ha-1), and then decreased under high N level. As N level increased, the ash, crude protein and amylose content varied from 1.36 to 2.25 %, from 7.99 to 15.84 % and from 22.69 to 27.64 %, respectively. The gelatinization enthalpy significantly increased with the range of 3.46-5.66 J/g, while no change was found in crystalline structure of common buckwheat flour. These results indicated that appropriate N application could effectively improve the endosperm development, starch synthesis and accumulation, and grain properties of common buckwheat, with the best effect under the level of 180 kg N ha-1.

Extraction and characterization of waxy and normal barley ß-glucans and their effects on waxy and normal barley starch pasting and degradation properties and mash filtration rate.

Interactions between ß-glucan and starch influence the health benefits of barley-based foods and barley brewing performance. Here, we characterized ß-glucans from waxy and normal barley varieties and compared the effects of different ß-glucans on the pasting and degradation of waxy and normal barley starches as well as the filterability of mashes from unmalted waxy and normal barley. Waxy barley Zangqing18 ß-glucan displayed more compact micrographic features, higher molecular weight, larger particle size, higher thermal decomposition temperature and lower rheological viscosity than normal barley Zangqing2000 ß-glucan. ß-Glucan not only significantly decreased the pasting viscosities of waxy and normal starches but also lowered the pasting temperatures and peak times of normal starch, likely by inhibiting granule swelling and disrupting the integrity of the continuous phase. ß-Glucan also decreased in vitro digestion extent of starch and increased the resistant starch. The unmalted waxy barley had a mash filtration rate much faster than normal barley because starch and ß-glucan in waxy barley were rapidly and completely digested and formed more open filter passages. The effects of ß-glucan on starch properties varied with the types and contents of ß-glucans, whilst the types of starches showed more significant effects. CHEMICAL COMPOUNDS STUDIED: ß-Glucan (Pubchem CID: 439262); Amylopectin (Pubchem CID: 439207); Starch (Pubchem CID: 156595876).

Anthocyanins from Aronia melanocarpa Bound to Amylopectin Nanoparticles: Tissue Distribution and In Vivo Oxidative Damage Protection.

The in vivo applications of anthocyanins are limited by their instability. Nano-encapsulation using amylopectin nanoparticles (APNPs) stabilizes anthocyanins to deliver them to tissues to ameliorate their physiological functions. Herein, rats are fed four Aronia melanocarpa anthocyanins encapsulated with APNPs, and their subsequent distributions and bioactivity in nine tissues are revealed using UHPLC-MS. Among digestive tissues, the concentration of the APNP-protected cyanidin 3-O-arabinoside in the stomach is 134.54% of that of the free anthocyanin, while among non-digestive tissues, the APNP-protected cyanidin 3-O-glucoside concentration in the lungs improved by 125.49%. Concentration maxima "double peaks" in the liver and kidney arise from different modes of transport. Sustained release of anthocyanins from anthocyanin-APNPs and stable concentration curves suggest controlled delivery, with most APNPs consumed in the digestive system. APNPs did not affect the overall anthocyanin absorption time or tissues. The superoxide dismutase and malondialdehyde concentrations indicate that APNPs enhance the oxidative damage protection in vivo.

Identification of a new allele of soluble starch synthase IIIa involved in the elongation of amylopectin long chains in a chalky rice mutant.

A chalky endosperm mutant (GM03) induced from an indica rice GLA4 was used to investigate the functional gene in starch biosynthesis. Bulked segregant analysis and sanger sequencing determined that a novel mutation in soluble starch synthase IIIa (SSIIIa) is responsible for the chalky phenotype in GM03. Complementary test by transforming the active SSIIIa gene driven by its native promoter to GM03 recovered the phenotype to its wildtype. The expression of SSIIIa was significantly decreased, while SSIIIa protein was not detected in GM03. The mutation of SSIIIa led to increased expression of most of starch synthesis related genes and elevated the levels of most of proteins in GM03. The CRISPR/Cas9 technology was used for targeted disruption of SSIIIa, and the mutant lines exhibited chalky endosperm which phenocopied the GM03. Additionally, the starch fine structure in the knockout mutant lines ss3a-1 and ss3a-2 was similar with the GM03, which showed increased amylose content, higher proportions of B1 and B2 chains, much lower proportions of B3 chains and decreased degree of crystallinity, leading to altered thermal properties with lower gelatinization temperature and enthalpy. Collectively, these results suggested that SSIIIa plays an important role in starch synthesis by elongating amylopectin long chains in rice.

Insights into the Supramolecular Structure and Degradation Mechanisms of Starch from Different Botanical Sources as Affected by Extrusion-based 3D Printing.

Extrusion-based 3D printing has emerged as the most versatile additive manufacturing technique for the printing of practically any material. However, 3D printing of functional materials often activates thermo-mechanical degradation, which affects the 3D shape quality. Herein, we describe the structural changes of eight different starch sources (normal or waxy) as a consequence of the temperature of an extrusion-based 3D printing system through in-depth characterization of their molecular and structural changes. The combination of size-exclusion chromatography, small-angle X-ray scattering, X-ray diffraction, dynamic viscoelasticity measurements, and in vitro digestion has offered an extensive picture of the structural and biological transformations of starch varieties. Depending on the 3D printing conditions, either gelatinization was attained ("moderate" condition) or single-amylose helix formation was induced ("extreme" condition). The stiff amylopectin crystallites in starch granules were more susceptible to thermo-mechanical degradation compared to flexible amorphous amylose. The crystalline morphology of the starch varieties varied from B-type crystallinity for the starch 3D printing at the "moderate" condition to a mixture of C- and V-type crystallinity regarding the "extreme" condition. The "extreme" condition reduced the viscoelasticity of 3D-printed starches but increased the starch digestibility rate/extent. In contrast, the "moderate" condition increased the viscoelastic moduli, decreasing the starch digestion rate/extent. This was more considerable mainly regarding the waxy starch varieties. Finally, normal starch varieties presented a well-defined shape fidelity, being able to form a stable structure, whereas waxy starches exhibited a non-well-defined structure and were not able to maintain their integrity after printing. The results of this research allow us to monitor the degradability of a variety of starch cultivars to create starch-based 3D structures, in which the local structure can be controlled based on the 3D printing parameters.

Maternal haploid waxy maize, environmental effects on grain yield, and quality parameters.

Waxy maize (Zea mays L. var. ceratina Kulesh) is used in the food and textile sectors, amylopectin has an important place in the adhesive and paper sectors as well. These sectors have to buy waxy maize from abroad because there is no waxy maize variety registered yet in Turkey. In vivo maternal haploid technique was applied to obtain doubled haploid (DH) waxy lines in a short time. RWS, RWK-76, and their hybrid RWS × RWK-76 maternal haploid inducers were used as male parents in vivo maternal haploid. SSRs markers were used to identify the genetic similarity between the number of 17 DH waxy lines. Similarity ratio ranged from 12% to 68% between DH waxy lines. DH waxy lines were used in crossbreeding and created 24 hybrids. Iodine tests were made on DH waxy lines and their hybrids and analyzed some quality parameters of hybrids. Candidate waxy hybrids were selected from the progeny nursery trial. Several 16 waxy and 3 check hybrids were experimented within three locations and the average grain yield of waxy and check hybrids ranged from 8.4-12.7 t/ha and 12.7-16.2 t/ha respectively. PCA biplot analysis using the data of the average of three locations and genotype × environment interaction was determined. PC1 and PC2 variation percentages were found to be 18.32% and 75.22%, respectively. ADAX-14, ADAX-13R, ADAX-13, and ADAX-19 waxy varieties are more stable in terms of yield than other hybrids. The difference between varieties was found statistically significant for protein, oil, starch, hectoliter, and 1000 grain weight.

Further slowing down of hydrolysis of amylose heated with black soybean extract by treating with nitrite under gastric conditions.

Black soybean (BSB), which contains cyanidin-3-O-glucoside (C3G) and procyanidins, is cooked with rice in Japan. The color of the cooked rice is purplish red due to the binding of C3G and reddish oxidation products of procyanidins. These components can slowdown pancreatin-induced hydrolysis of amylose more significantly than the hydrolysis of amylopectin, and can react with nitrous acid in the stomach. This manuscript deals with the effects of nitrous acid on pancreatin-induced hydrolysis of amylose heated with BSB extract. The hydrolysis of amylose heated with BSB extract was slow, and the slowdown was due to the binding of C3G/its degradation products and degradation products of procyanidins. The amylose hydrolysis was slowed down further by treating with nitrite under gastric conditions. The further slowdown was discussed to be due to the binding of the products, which were formed by the reaction of procyanidins with nitrous acid, to amylose. In the products, dinitroprocyanidins were included. In this way, the digestibility of amylose heated with BSB extract can be slowed down further by reacting with nitrous acid in the stomach.

Mutation of TL1, encoding a novel C2H2 zinc finger protein, improves grains eating and cooking quality in rice.

KEY MESSAGE: The cloning and characterization of a novel C2H2 zinc finger protein that affects rice eating and cooking quality by regulating amylose content and amylopectin chain-length distribution in rice. One of the major objectives in rice breeding aims to increase simultaneously yield and grain quality especially eating and cooking quality (ECQ). Controlling amylose content (AC) and amylopectin chain-length distribution (ACLD) in rice is a major strategy for improving rice ECQ. Previous studies show that some starch synthesis-related genes (SSRGs) are required for normal AC and ACLD, but its underlying regulating network is still unclear. Here, we report the cloning and characterization of a novel C2H2 zinc finger protein TL1 (Translucent endosperm 1) that positively regulates amylose synthesis in rice grains. Loss of TL1 function reduced apparent amylose content (AAC), total starch, gel consistency, and gelatinisation temperature, whereas increased viscosity, total lipid, and ratio of amylopectin A chains with degree of polymerization (DP) 6-12 to B1 chains with DP 13-24, resulting in an enhanced grain ECQ. The improved ECQ was accompanied by altered expression patterns of several tested SSRGs in tl1 mutant grains. Furthermore, knockout of TL1 in the high-yielding rice variety JiaHua NO.1 reduced AAC without obvious side effects on major agronomic traits. These findings expand our understanding of the regulating networks of grain starch metabolism and provide new insights into how rice ECQ quality can be improved via genetic approach.