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Summary: It is well known that the overall quality of japonica/geng rice is superior to that of indica/xian rice varieties. However, the molecular mechanisms underlying the quality disparities between these two subspecies of rice are still largely unknown. In this study, we have pinpointed a gene homologous to SLR1, termed LCG1, exhibiting significant expression during early caryopsis development and playing a specific role in regulating rice chalkiness and taste by affecting the accumulation of grain storage components, starch granule structure and chain length distribution of amylopectin. LCG1 physically interacts with OsBP5 and indirectly influences the expression of the amylose synthesis gene Waxy (Wx) by hindering the transcriptional activity of the OsBP5/OsEBP89 complex. Notably, sequence variations in the promoter region of LCG1 result in enhanced transcription in japonica rice accessions. This leads to elevated LCG1 expression in CSSL‐LCG1Nip, thereby enhancing rice quality. Our research elucidates the molecular mechanism underlying the impact of the LCG1‐OsBP5/OsEBP89‐Wx regulatory pathway on rice chalkiness and taste quality, offering new genetic resources for improving the indica rice quality. [ABSTRACT FROM AUTHOR]

Amylose content, the key determinant of rice eating and cooking quality, is regulated primarily by the Waxy (Wx) gene. We adjusted the amylose content and transparency of semi-glutinous japonica rice carrying the Wxmp allele by genome editing of upstream open reading frame 6 (uORF6) of Wx.

To explore the molecular mechanism behind maize grain quality and use of different gene stacking to improve the nutritional quality of grain, marker-assisted selection (MAS) was used to select three recessive mutant lines containing o2o16wx, along with the double-recessive mutant lines containing o2o16, o2wx, and o16wx. The resulting seeds were taken for transcriptome sequencing analysis 18 days after pollination (DAP). Results: Compared with the recurrent parent genes, in the lysine synthesis pathway, the gene pyramiding lines (o2o16wx, o2wx, and o16wx) revealed that the gene encoding aspartate kinase (AK) was up-regulated and promoted lysine synthesis. In the lysine degradation pathway, 'QCL8010_1' (o2o16wx) revealed that the gene encoding saccharopine dehydrogenase (LKR/SDH) was down-regulated. In addition, the gene pyramiding lines (o2o16wx, o2o16, and o16wx) indicated that the gene encoding 2-oxoglutarate dehydrogenase E1 component (OGDH) was down-regulated, inhibiting the degradation of lysine. In the tryptophan synthesis pathway, the genes encoding anthranilate synthase (AS), anthranilate synthase (APT), and tryptophan synthase (TS) were up-regulated (in o2o16wx, o2o16, o2wx, and o16wx), and promote tryptophan synthesis. In the tryptophan degradation pathway, it was revealed that the genes encoding indole-3-producing oxidase (IAAO) and indole-3-pyruvate monooxygenase (YUCCA) were down-regulated. These results provide a reference for revealing the mechanism of the o2, o16, and wx with different gene pyramiding to improve grain quality in maize. [ABSTRACT FROM AUTHOR]

Amylose content (AC) is a crucial determinant of the eating and cooking quality (ECQ) of rice, with low AC varieties exhibiting a softer texture and greater stickiness -attributes that enhance palatability and are desirable in specific culinary contexts. To harness these traits, significant efforts have been made to manipulate AC to improve rice ECQ. Our research utilized the MutMap+ approach to identify LAC6/TL1, a gene that is an allele of Du13, responsible for low AC. LAC6 encodes a C2H2 zinc finger protein, which specifically increases the splicing efficiency of the Wxb allele without affecting the Wxa allele. Functional studies of LAC6 revealed that its proper integration could rectify the undesirable AC phenotype, whereas mutations within this gene led to reduced AC and were associated with shorter grain length and decreased thousand-grain weight. Despite these drawbacks, such mutations positively impact rice palatability, presenting a trade-off between grain size and eating quality. To address the challenges posed by the reduced grain weight associated with LAC6 mutations, we developed a specific molecular marker for LAC6, which has been effectively used in breeding programs to select lac6/tl1/du13 homozygous individuals with larger grain size. Our findings demonstrate that the "small grain" trait associated with lac6/tl1/du13 can be effectively mitigated through combined phenotype-based and marker-assisted selection. This study highlights the potential of lac6/tl1/du13 as a valuable gene for breeding novel, high-quality soft rice varieties through targeted breeding strategies. [ABSTRACT FROM AUTHOR]

Amylose content (AC) stands as a pivotal determinant of rice grain quality, primarily governed by the Waxy gene (Wx). The allelic variation within this gene, particularly the presence of the Wxmp allele derived from the ancestral Wxmq allele, significantly influences AC and is prevalent among soft japonica rice varieties in southern China. Although both alleles are associated with lower AC, there remains a paucity of detailed understanding regarding the interplay between specific functional single nucleotide polymorphisms (SNPs) within these alleles and the overarching rice grain quality. To investigate this, we engineered three distinct transgenic rice lines, each harboring the Wxmp, Wxmq, or Wxb−5c alleles in the background of the glutinous rice cultivar Nip(wx). This suite of transgenic rice lines showcased varying degrees of grain transparency inversely correlated to AC, which in turn influenced other physicochemical properties of the rice grains, such as taste value of cooked rice, gel consistency, and starch pasting properties. Additionally, analyses of gene expression and enzyme activity revealed that the functional SNPs, Ex4-53G to A and Ex5-53T to C, lead to a decline in the activity of granule-bound starch synthase I (GBSSI) without altering expression levels. [ABSTRACT FROM AUTHOR]

Rice eating and cooking quality (ECQ) is one of the most important qualities and it has attracted more and more interests of consumers and breeders. SSIIa and Wx are the major genes affecting rice ECQ, and several alleles of SSIIa and Wx have been identified. In this study, the effects of different allele combinations of SSIIa and Wx on rice ECQ were explored by analyzing amylose content (AC), gelatinization temperature (GT), and Rapid Viscosity Analyzer (RVA) profile properties of one natural population which is composed of 30 glutinous rice varieties and 342 non-glutinous varieties. Nine allele combinations of SSIIa and Wx were identified in all the non-glutinous varieties. The allele combination of SSIIac/Wxb harbored higher BDV, lower SBV, intermediate AC, and higher GT, which means better taste quality but more cooking time. For non-glutinous rice, stepwise regression analysis showed that RVA profile properties are mainly controlled by polymorphic loci of Wx-Int1(G/T) and Wx-EX10(C/T); AC, PV, BDV, and SBV are mainly controlled by Wx-Int1(G/T); HPV, CPV, and CSV are mainly controlled by Wx-EX10(C/T); GT is mainly controlled by SSIIa-EX8(GC/TT). Regression equation models also confirmed that SSIIac/Wxb had better ECQ compared to other allele combinations, indicating its potential to improve rice quality. Our results may provide useful clues for elite alleles of SSIIa and Wx for introgression to other rice varieties to improve rice ECQ. [ABSTRACT FROM AUTHOR]

Wx is the key gene that controls amylose content (AC), and various alleles have been found in rice populations. Wxb is the major allele in japonica and produces moderate AC (15~18%). It was recently found that editing the promoter of Wx could produce a series of alleles that have different Wx activities. Although some studies have edited the promoter, few studies have focused on the natural variations in Wx. Here, we used the Rice3K database to investigate variations in the Wx promoter and found that the allele Wx1764178 (A/G) has a higher LD (linkage disequilibrium) with the two key SNPs (1765751, T/G; 1768006, A/C), which could produce different Wx alleles and influence AC, as reported previously. Further study showed that the Wx1764178 allele (A/G) is functional and influences the expression of Wx positively. Editing the A allele using CRISPR‒Cas9 produced 36 and 3 bp deletions and caused a decrease in the expression of Wx. The apparent amylose content (AAC) in the edited lines was decreased by 7.09% and 11.50% compared with that of the wild type, which was the japonica variety Nipponbare with Wxb and the A allele at 1764178, while a complementary line with the G allele showed a lower AAC than the A allele with no effect on other agronomic traits. The AAC of the edited lines showed a higher increase than that of the wild type (Nipponbare, Wxb) in low-nitrogen conditions relative to high-nitrogen conditions. We also developed a dCAPS marker to identify the allele and found that the G allele has widely been used (82.95%) in japonica-bred varieties from Jiangsu Province, China. Overall, we found a functional allele (Wx1764178, A/G) in the Wx promoter that could affect AAC in japonica cultivars and be developed as markers for quality improvement in rice breeding programs. [ABSTRACT FROM AUTHOR]

In rice cultivation, the traits of semi-dwarfism and glutinous texture are pivotal for optimizing yield potential and grain quality, respectively. Xiangdaowan (XDW) rice, renowned for its exceptional aromatic properties, has faced challenges due to its tall stature and high amylose content, resulting in poor lodging resistance and suboptimal culinary attributes. To address these issues, we employed CRISPR/Cas9 technology to precisely edit the SD1 and Wx genes in XDW rice, leading to the development of stable genetically homozygous lines with desired semi-dwarf and glutinous characteristics. The sd1-wx mutant lines exhibited reduced gibberellin content, plant height, and amylose content, while maintaining hardly changed germination rate and other key agronomic traits. Importantly, our study demonstrated that exogenous GA3 application effectively promoted growth by compensating for the deficiency of endogenous gibberellin. Based on this, a semi-dwarf glutinous elite rice (Oryza sativa L.) Lines was developed without too much effect on most agronomic traits. Furthermore, a comparative transcriptome analysis unveiled that differentially expressed genes (DEGs) were primarily associated with the anchored component of the membrane, hydrogen peroxide catabolic process, peroxidase activity, terpene synthase activity, and apoplast. Additionally, terpene synthase genes involved in catalyzing the biosynthesis of diterpenoids to gibberellins were enriched and significantly down-regulated. This comprehensive study provides an efficient method for simultaneously enhancing rice plant height and quality, paving the way for the development of lodging-resistant and high-quality rice varieties. [ABSTRACT FROM AUTHOR]

Amylose content (AC) stands as a pivotal determinant of rice grain quality, primarily governed by the Waxy gene (Wx). The allelic variation within this gene, particularly the presence of the Wxmp allele derived from the ancestral Wxmq allele, significantly influences AC and is prevalent among soft japonica rice varieties in southern China. Although both alleles are associated with lower AC, there remains a paucity of detailed understanding regarding the interplay between specific functional single nucleotide polymorphisms (SNPs) within these alleles and the overarching rice grain quality. To investigate this, we engineered three distinct transgenic rice lines, each harboring the Wxmp, Wxmq, or Wxb−5c alleles in the background of the glutinous rice cultivar Nip(wx). This suite of transgenic rice lines showcased varying degrees of grain transparency inversely correlated to AC, which in turn influenced other physicochemical properties of the rice grains, such as taste value of cooked rice, gel consistency, and starch pasting properties. Additionally, analyses of gene expression and enzyme activity revealed that the functional SNPs, Ex4-53G to A and Ex5-53T to C, lead to a decline in the activity of granule-bound starch synthase I (GBSSI) without altering expression levels.

Grain chalkiness is an important index of rice appearance quality and is negatively associated with rice processing and eating quality. However, the genetic mechanism underlying chalkiness formation is largely unknown. To identify the genetic basis of chalkiness, 410 recombinant inbred lines (RILs) derived from two representative indica rice varieties, Shuhui498 (R498) and Yihui3551 (R3551), were used to discover quantitative trait loci (QTLs). The two parental lines and RILs were grown in three locations in China under three controlled fertilizer application levels. Analyses indicated that chalkiness was significantly affected by genotype, the environment, and the interaction between the two, and that heritability was high. Several QTLs were isolated, including the two stable QTLs qCGP6 and qCGP8. Fine mapping and candidate gene verification of qCGP6 showed that Wx may play a key role in chalkiness formation. Chromosomal segment substitution lines (CSSLs) and near-isogenic lines (NILs) carrying the Wx a or Wx in allele produced more chalky grain than the R498 parent. A similar result was also observed in the 3611 background. Notably, the effect of the Wx genotype on rice chalkiness was shown to be dependent on environmental conditions, and Wx alleles exhibited different sensitivities to shading treatment. Using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9), the Wx a promoter region was successfully edited; down-regulating Wx alleviates chalkiness formation in NILR498- Wx a . This study developed a new strategy for synergistic improvement of eating and appearance qualities in rice, and created a novel Wx allele with great potential in breeding applications. [ABSTRACT FROM AUTHOR]

MicroRNA transcriptomes from fresh tumors and the adjacent normal tissues were profiled in 10 Korean patients diagnosed with lung adenocarcinoma using a next-generation sequencing (NGS) technique called miRNA-seq. The sequencing quality was assessed using FastQC, and low-quality or adapter-contaminated portions of the reads were removed using Trim Galore. Quality-assured reads were analyzed using miRDeep2 and Bowtie. The abundance of known miRNAs was estimated using the reads per million (RPM) normalization method. Subsequently, using DESeq2 and Wx, we identified differentially expressed miRNAs and potential miRNA biomarkers for lung adenocarcinoma tissues compared to adjacent normal tissues, respectively. We defined reliable miRNA biomarkers for lung adenocarcinoma as those detected by both methods. The miRNA-seq data are available in the Gene Expression Omnibus (GEO) database under accession number GSE196633, and all processed data can be accessed via the Mendeley data website. Dataset:  https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE196633 and https://data.mendeley.com/datasets/vp977psjcb/2. Dataset License: CC0 [ABSTRACT FROM AUTHOR]

Wx is the key gene that controls amylose content (AC), and various alleles have been found in rice populations. Wxb is the major allele in japonica and produces moderate AC (15~18%). It was recently found that editing the promoter of Wx could produce a series of alleles that have different Wx activities. Although some studies have edited the promoter, few studies have focused on the natural variations in Wx. Here, we used the Rice3K database to investigate variations in the Wx promoter and found that the allele Wx1764178 (A/G) has a higher LD (linkage disequilibrium) with the two key SNPs (1765751, T/G; 1768006, A/C), which could produce different Wx alleles and influence AC, as reported previously. Further study showed that the Wx1764178 allele (A/G) is functional and influences the expression of Wx positively. Editing the A allele using CRISPR‒Cas9 produced 36 and 3 bp deletions and caused a decrease in the expression of Wx. The apparent amylose content (AAC) in the edited lines was decreased by 7.09% and 11.50% compared with that of the wild type, which was the japonica variety Nipponbare with Wxb and the A allele at 1764178, while a complementary line with the G allele showed a lower AAC than the A allele with no effect on other agronomic traits. The AAC of the edited lines showed a higher increase than that of the wild type (Nipponbare, Wxb) in low-nitrogen conditions relative to high-nitrogen conditions. We also developed a dCAPS marker to identify the allele and found that the G allele has widely been used (82.95%) in japonica-bred varieties from Jiangsu Province, China. Overall, we found a functional allele (Wx1764178, A/G) in the Wx promoter that could affect AAC in japonica cultivars and be developed as markers for quality improvement in rice breeding programs.

Global food security has benefited from the development and promotion of the two-line hybrid rice system. Excellent eating quality determines the market competitiveness of hybrid rice varieties based on achieving the fundamental requirements of high yield and good adaptability. Developing sterile and restorer lines with improved quality for two-line hybrid breeding by editing quality genes with clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 is an efficient and practical alternative to the lengthy and laborious process of conventional breeding to improve rice quality. We edited Wx and OsBADH2 using CRISPR/Cas9 technology to produce both homozygous male sterile mutant lines and homozygous restorer mutant lines with Cas9-free. These mutants have a much lower amylose content while having a significantly higher 2-acetyl-1-pyrroline aroma content. Based on this, a fragrant glutinous hybrid rice was developed without too much effect on most agronomic traits. This study demonstrates the use of CRISPR/Cas9 in creating two-line fragrant glutinous hybrid rice by editing the components of the male sterile and the restorative lines. [ABSTRACT FROM AUTHOR]

The improvement of the cooking and eating quality (CEQ) of rice is one of the major objectives of current rice-breeding programs. A few major genes such as Waxy (Wx) and starch synthase IIa (SSIIa) have been successfully applied in molecular breeding. However, their interactive effects on CEQ have not been fully understood. In this study, a recombinant inbred line (RIL) population was constructed by crossing the white-core mutant GM645 with the transparent phenotype of the japonica rice variety Tainung 67 (TN67). GM645 and TN67 contain different alleles of FLOURY ENDOSPERM2 (FLO2), Wx, and SSIIa. The effects of different allele combinations of FLO2, Wx, and SSIIa on the CEQ of rice were investigated. The inbred lines with the mutation allele flo2 had a significantly lower apparent amylose content (AAC), viscosity characteristics except for setback (SB), and gel texture properties compared to those lines with the FLO2 allele. The allelic combination of FLO2 and Wx significantly affected the AAC, breakdown (BD), and gel textural properties, which could explain most of the variations in those rice quality traits that were correlated with AAC. The allelic combination of FLO2 and SSIIa significantly affected the hot paste viscosity (HPV) and pasting temperature (PT). The Wx × SSIIa interaction had a significant effect on the PT. The interaction of FLO2, Wx and SSIIa significantly affected the AAC, cold paste viscosity (CPV), PT, and consistency viscosity (CS). These results highlight the important roles of these quality-related genes in regulating the CEQ of rice and provide new clues for rice-quality improvement by marker-assisted selection. [ABSTRACT FROM AUTHOR]

MicroRNA transcriptomes from fresh tumors and the adjacent normal tissues were profiled in 10 Korean patients diagnosed with lung adenocarcinoma using a next-generation sequencing (NGS) technique called miRNA-seq. The sequencing quality was assessed using FastQC, and low-quality or adapter-contaminated portions of the reads were removed using Trim Galore. Quality-assured reads were analyzed using miRDeep2 and Bowtie. The abundance of known miRNAs was estimated using the reads per million (RPM) normalization method. Subsequently, using DESeq2 and Wx, we identified differentially expressed miRNAs and potential miRNA biomarkers for lung adenocarcinoma tissues compared to adjacent normal tissues, respectively. We defined reliable miRNA biomarkers for lung adenocarcinoma as those detected by both methods. The miRNA-seq data are available in the Gene Expression Omnibus (GEO) database under accession number GSE196633, and all processed data can be accessed via the Mendeley data website.

Amylose largely determines rice grain quality profiles. The process of rice amylose biosynthesis is mainly driven by the waxy (Wx) gene, which also affects the diversity of amylose content. The present study assessed the grain quality profiles, starch fine structure, and crystallinity characteristics of the near-isogenic lines Q11(Wxlv), NIL(Wxa), and NIL(Wxb) in the indica rice Q11 background containing different Wx alleles. Q11(Wxlv) rice contained a relatively higher amylose level but very soft gel consistency and low starch viscosity, compared with rice lines carrying Wxa and Wxb. In addition, starch fine structure analysis revealed a remarkable decrease in the relative area ratio of the short amylopectin fraction but an increased amylose fraction in Q11(Wxlv) rice. Chain length distribution analysis showed that Q11(Wxlv) rice contained less amylopectin short chains but more intermediate chains, which decreased the crystallinity and lamellar peak intensity, compared with those of NIL(Wxa) and NIL(Wxb) rice. Additionally, the starches in developing grains showed different accumulation profiles among the three rice lines. Moreover, significant differences in starch gelatinization and retrogradation characteristics were observed between near-isogenic lines, which were caused by variation in starch fine structure. These findings revealed the effects of Wxlv on rice grain quality and the fine structure of starch in indica rice. [ABSTRACT FROM AUTHOR]

Amylose content (AC), which is regulated by the Waxy (Wx) gene, is a major indicator of eating and cooking quality (ECQ) in rice (Oryza sativa). Thus far, only a limited number of mutations in the N‐terminal domain of Wx were found to have a major impact on the AC of rice grains and no mutations with such effects were reported for other regions of the Wx protein. Here, nucleotide substitutions in the middle region of Wx were generated by adenine and cytosine base editors. The nucleotide substitutions led to changes in 15 amino acid residues of Wx, and a series of novel Wx alleles with ACs of 0.3%–29.43% (wild type with AC of 19.87%) were obtained. Importantly, the waxyabe2 allele showed a "soft rice" AC, improved ECQ, favorable appearance, and no undesirable agronomic traits. The transgenes were removed from the waxyabe2 progeny, generating a promising breeding material for improving rice grain quality. [ABSTRACT FROM AUTHOR]

The improvement of the cooking and eating quality (CEQ) of rice is one of the major objectives of current rice-breeding programs. A few major genes such as Waxy (Wx) and starch synthase IIa (SSIIa) have been successfully applied in molecular breeding. However, their interactive effects on CEQ have not been fully understood. In this study, a recombinant inbred line (RIL) population was constructed by crossing the white-core mutant GM645 with the transparent phenotype of the japonica rice variety Tainung 67 (TN67). GM645 and TN67 contain different alleles of FLOURY ENDOSPERM2 (FLO2), Wx, and SSIIa. The effects of different allele combinations of FLO2, Wx, and SSIIa on the CEQ of rice were investigated. The inbred lines with the mutation allele flo2 had a significantly lower apparent amylose content (AAC), viscosity characteristics except for setback (SB), and gel texture properties compared to those lines with the FLO2 allele. The allelic combination of FLO2 and Wx significantly affected the AAC, breakdown (BD), and gel textural properties, which could explain most of the variations in those rice quality traits that were correlated with AAC. The allelic combination of FLO2 and SSIIa significantly affected the hot paste viscosity (HPV) and pasting temperature (PT). The Wx × SSIIa interaction had a significant effect on the PT. The interaction of FLO2, Wx and SSIIa significantly affected the AAC, cold paste viscosity (CPV), PT, and consistency viscosity (CS). These results highlight the important roles of these quality-related genes in regulating the CEQ of rice and provide new clues for rice-quality improvement by marker-assisted selection.

Key Message: Coordinated regulation of amylose and amylopectin synthesis via manipulation of SSII-2, SSII-3 and Wx expression in endosperm can improve rice eating and cooking quality. With increasing rice consumption worldwide, many researchers are working to increase the yield and improve grain quality, especially eating and cooking quality (ECQ). The rice ECQ is mainly controlled by the expression of starch synthesis-related genes (SSRGs) in endosperm. Although the Wx and SSII-3/SSIIa/ALK genes, two major SSRGs, have been manipulated to improve rice ECQ via various breeding approaches, new methods to further improve ECQ are desired. In our previous study, we enhanced rice ECQ by knocking down SSII-2 expression in the japonica Nipponbare cultivar (carrying the Wxb allele) via RNA interference. Herein, the SSII-2 RNAi was introduced into two Nipponbare-derived near-isogenic lines (NILs), Nip(Wxa) and Nip(wx), carrying Wxa and wx alleles respond for high and no amylose levels, respectively. Analysis of physicochemical properties revealed that the improved grain quality of SSII-2 RNAi transgenic lines was achieved by coordinated downregulating the expression of SSII-2, SSII-3 and Wx. To further confirm this conclusion, we generated ssii-2, ssii-3 and ssii-2ssii-3 mutants via CRISPR/Cas9 technique. The amylopectin structure of the resulting ssii-2sii-3 mutants was similar to that in SSII-2 RNAi transgenic lines, and the absence of SSII-2 decreased the amylose content, gelatinisation temperature and rapid visco-analyser profile, indicating essential roles for SSII-2 in the regulation of amylopectin biosynthesis and amylose content in rice endosperm. The effect of SSII-2 was seen only when the activity of SSII-3 was very low or lacking. Our study provides novel approaches and valuable germplasm resources for improving ECQ via plant breeding. [ABSTRACT FROM AUTHOR]

According to geographic origin and amylose content, 87 investigated japonica rice cultivars were divided into three types: Jiangsu common japonica rice, Northeast japonica rice and Jiangsu semi-waxy japonica rice. In order to explore the eating quality characteristics of the three types of japonica rice and their molecular basis, the physicochemical properties and the sequence of major genes Wx and OsSSIIa for starch synthesis were analyzed. Compared with Northeast japonica rice, Jiangsu common japonica rice had lower amylose content, higher protein content and gelatinization temperature, and similar gel consistency and RVA profiles. Compared with Northeast japonica rice and Jiangsu common japonica rice, Jiangsu semi-waxy japonica rice contained the lowest amylose content, the highest gel consistency and unique RVA profile. Variation coefficients of all characters except for setback value were small in three types of rice varieties, indicating that there was a great range of retrogradation within all the cultivars. The correlation coefficients among traits were different among three types of japonica rice, which indicated that the correlations were affected by the ecological conditions of different regions and properties of varieties. By cluster analysis, 87 japonica rice varieties (lines) were divided into four groups. Jiangsu common and Northeast japonica rice varieties (lines) were intersected in group I and group II. Group III was composed of two lines of Jiangsu common japonica rice and one semi-waxy variety (Xudao 9), and the most of their character values were ranged between those common japonica rice and semi-waxy japonica rice. The Jiangsu semi-waxy rice varieties (lines) with unique physicochemical characteristics were clustered into group IV. By sequence analysis, there was no difference in Wxb gene between Jiangsu common japonica rice and Northeast japonica rice, but in Jiangsu semi-waxy japonica rice G/A substitution occurred in Exon 4 of Wxb, which is the Wxmp allele. The higher gelatinization temperature in some of Jiangsu japonica rice varieties (lines) was controlled by the allele of OsSSIIa (G-GC). [ABSTRACT FROM AUTHOR]

Amylose largely determines rice grain quality profiles. The process of rice amylose biosynthesis is mainly driven by the waxy (Wx) gene, which also affects the diversity of amylose content. The present study assessed the grain quality profiles, starch fine structure, and crystallinity characteristics of the near-isogenic lines Q11(Wxlv), NIL(Wxa), and NIL(Wxb) in the indica rice Q11 background containing different Wx alleles. Q11(Wxlv) rice contained a relatively higher amylose level but very soft gel consistency and low starch viscosity, compared with rice lines carrying Wxa and Wxb. In addition, starch fine structure analysis revealed a remarkable decrease in the relative area ratio of the short amylopectin fraction but an increased amylose fraction in Q11(Wxlv) rice. Chain length distribution analysis showed that Q11(Wxlv) rice contained less amylopectin short chains but more intermediate chains, which decreased the crystallinity and lamellar peak intensity, compared with those of NIL(Wxa) and NIL(Wxb) rice. Additionally, the starches in developing grains showed different accumulation profiles among the three rice lines. Moreover, significant differences in starch gelatinization and retrogradation characteristics were observed between near-isogenic lines, which were caused by variation in starch fine structure. These findings revealed the effects of Wxlv on rice grain quality and the fine structure of starch in indica rice.

In rice cultivation, the traits of semi-dwarfism and glutinous texture are pivotal for optimizing yield potential and grain quality, respectively. Xiangdaowan (XDW) rice, renowned for its exceptional aromatic properties, has faced challenges due to its tall stature and high amylose content, resulting in poor lodging resistance and suboptimal culinary attributes. To address these issues, we employed CRISPR/Cas9 technology to precisely edit the SD1 and Wx genes in XDW rice, leading to the development of stable genetically homozygous lines with desired semi-dwarf and glutinous characteristics. The sd1-wx mutant lines exhibited reduced gibberellin content, plant height, and amylose content, while maintaining hardly changed germination rate and other key agronomic traits. Importantly, our study demonstrated that exogenous GA 3 application effectively promoted growth by compensating for the deficiency of endogenous gibberellin. Based on this, a semi-dwarf glutinous elite rice ( Oryza sativa L.) Lines was developed without too much effect on most agronomic traits. Furthermore, a comparative transcriptome analysis unveiled that differentially expressed genes (DEGs) were primarily associated with the anchored component of the membrane, hydrogen peroxide catabolic process, peroxidase activity, terpene synthase activity, and apoplast. Additionally, terpene synthase genes involved in catalyzing the biosynthesis of diterpenoids to gibberellins were enriched and significantly down-regulated. This comprehensive study provides an efficient method for simultaneously enhancing rice plant height and quality, paving the way for the development of lodging-resistant and high-quality rice varieties., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Wang, Gao, Liu, Wang, Zhang, Wei, Lu, Li, Shi, Zhao, Zhou, Peng and Yuan.)

Abstract The waxy gene (Wx) in rice, which encodes the granule bound starch synthase enzyme, is responsible for amylose synthesis. Glutinous (sticky) rice has little or no amylose that can be used in various applications, such as brewing. In this study, knockout of the Wx gene with CRISPR/Cas9 technology was conducted in two elite japonica rice lines, Huaidao 5 (HD5) and Suken 118 (SK118), aiming to develop elite sticky rice varieties. We achieved six homozygous T 0 plants with more than 200 bp deletion in the Wx gene, as well as 36 wx -HD5 and 18 wx -SK118 homozygous transgene-free plants in the T 1 generation. The seeds of all the mutants were white and opaque, similar to those of sticky rice, and contained only 2.6%–3.2% amylose. Results of scanning electron microscopy showed that the quality of rice did not change. In conclusion, we successfully developed two elite sticky rice varieties. [ABSTRACT FROM AUTHOR]

Purpose. Reporting peculiarities of new forms of edible corn and prospects for their use in food production that have been created jointly by scientists from the Plant Production Institute nd. a. V. Ya. Yuryev of NAAS and Ustymiv­ka Experimental Station for Plant Production on the base of the environmentally friendly method that uses biochemical effects of natural mutations of corn grain endosperm structure. Methods. Field study, laboratory analysis, mathematical and statistical evaluation. Results. Original series of inbred lines as well as promising varieties and hyb­rids of sugar and waxy corns were obtained. Varieties and hybrids are universally usable for obtaining both fresh and canned products, characterized by good taste, provide yields of up to 10 tons per hectare of commercial products and aimed to grow in various soil and climatic conditions. Brief economic and morphological characteristics of corn varieties and hybrids created jointly by scientists from the Plant Production Institute nd. a. V. Ya. Yuryev of NAAS and Ustymivka Experimental Station for Plant Production were given. Conclusions. The use of biochemical effect of natural mutations of endosperm structure is cost-effective and environmentally friendly method for creating lines, varieties and hybrids of corn for direct consumption and processing into food products. ‘Biliavka’, ‘Nika’ varieties and ‘Binom F1’, ‘Solo F1’, ‘Twister F1’ hybrids created with the help of this method are promising for commercial use

The Wx gene encodes a granule-bound starch synthase (GBSS) and plays a key role in determining rice eating and cooking qualities (ECQs). SSIII-2 (SSIIIa), a member of the soluble starch synthases, is responsible for the synthesis of long chains of amylopectin. To investigate the effects of Wx and its interaction with SSIII-2 on grain ECQs, a population from a hybrid combination was established as a research material. The genotypes of SSIII-2 and the single nucleotide polymorphisms (SNPs) on intron1, exon6, and exon10 of Wx, and the physicochemical indicators and rapid visco analyzer (RVA) profile characteristics were analyzed. The results revealed various effects of SSIII-2 on rice quality under different backgrounds of Wx alleles. There was no obvious difference between different SSIII-2 alleles under the same background of Wxa, whereas there was a significant diversity under the same background of Wxb. Wxa had a dominant epistasis to SSIII-2 because the effect of SSIII-2 was masked by the massive synthesis of GBSS under Wxa. The apparent amylose content (AAC) was mainly controlled by the In1G/T SNP, and rice gel consistency (GC) was regulated by the Ex10C/T SNP. The combined effects of three SNPs had a significant influence on all ECQs and RVA profile parameters, except for gelatinization temperature. In1T-Ex6A-Ex10C and In1T-Ex6A-Ex10T were classified as being low AAC type. TT-AA-CC and TT-AA-TT had a low AAC and a soft GC. The combined effects of different SNPs of Wx are very important for rice quality breeding.

[Excerpt] For the fourth consecutive year, the Cornell Center for Real Estate and Finance and Cornell Real Estate Women hosted its annual WX Panel Discussion held at Statler Hall on March 11. WX New York Women Executives in Real Estate is an invitation-only association for New York's executive-level women actively engaged in the commercial real estate industry. Through networking opportunities and educational seminars, WX promotes the advancement of women in the industry for its diverse membership of owners, developers, brokers, architects, engineers, bankers, and lawyers. Sylvia Melikian (MBA '76) moderated the four member panel of distinguished WX executives who shared their experiences and strategies for achieving success in the industry and their respective fields of hospitality, banking, development, and asset management. Over forty undergraduate and graduate students in hotel administration, city and regional planning, real estate, and business attended the event. After the discussion, panelists and students enjoyed a networking reception in Statler Hall. Below are some highlights from the evening. [ABSTRACT FROM AUTHOR]

The Wx gene encodes a granule-bound starch synthase (GBSS) and plays a key role in determining rice eating and cooking qualities (ECQs). SSIII-2 (SSIIIa), a member of the soluble starch synthases, is responsible for the synthesis of long chains of amylopectin. To investigate the effects of Wx and its interaction with SSIII-2 on grain ECQs, a population from a hybrid combination was established as a research material. The genotypes of SSIII-2 and the single nucleotide polymorphisms (SNPs) on intron1, exon6, and exon10 of Wx, and the physicochemical indicators and rapid visco analyzer (RVA) profile characteristics were analyzed. The results revealed various effects of SSIII-2 on rice quality under different backgrounds of Wx alleles. There was no obvious difference between different SSIII-2 alleles under the same background of Wxa, whereas there was a significant diversity under the same background of Wxb. Wxa had a dominant epistasis to SSIII-2 because the effect of SSIII-2 was masked by the massive synthesis of GBSS under Wxa. The apparent amylose content (AAC) was mainly controlled by the In1G/T SNP, and rice gel consistency (GC) was regulated by the Ex10C/T SNP. The combined effects of three SNPs had a significant influence on all ECQs and RVA profile parameters, except for gelatinization temperature. In1T-Ex6A-Ex10C and In1T-Ex6A-Ex10T were classified as being low AAC type. TT-AA-CC and TT-AA-TT had a low AAC and a soft GC. The combined effects of different SNPs of Wx are very important for rice quality breeding. [ABSTRACT FROM AUTHOR]

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Development of high yielding and more palatable glutinous rice is an important goal in breeding and long-standing cultural interaction in Asia. In this study, the TGW6 and Wx, major genes conferring 1000 grain weight (GW) and amylose content (AC), were edited in a maintainer line by CRISPR/Cas9 technology. Four targets were assembled in pYLCRISPR/Cas9Pubi-H vector and T0 mutant plants were obtained through Agrobacterium mediated transformation with 90% mutation frequency having 28% homozygous mutations without off-target effects in three most likely sites of each target and expression level of target genes in mutant lines was significantly decreased (P < 0.01), the GW and gel consistency (GC) were increased, and the AC and gelatinization temperature (GT) were decreased significantly and grain appearance was opaque, while there was no change in starch content (SC) and other agronomic traits. Mutations were inheritable and some T1 plants were re-edited but T2 generation was completely stable. The pollen fertility status was randomly distributed, and the mutant maintainer lines were hybridized with Cytoplasmic Male Sterile (CMS) line 209A and after subsequent backcrossing the two glutinous CMS lines were obtained in BC2F1. The identified proteins from anthers of CMS and maintainer line were closely associated with transcription, metabolism, signal transduction, and protein biosynthesis. Putative mitochondrial NAD+-dependent malic enzyme was absent in CMS line which caused the pollen sterility because of insufficient energy, while upregulation of putative acetyl-CoA synthetase and Isoamylase in both lines might have strong relationship with CMS and amylose content. High yielding glutinous CMS lines will facilitate hybrid rice breeding and investigations of proteins linked to male sterility will provide the insights to complicated metabolic network in anther development.

A glutinous texture of endosperm is one of the important traits of rice (Oyza sativa L.). Northern Laos is known as a center of glutinous rice diversity. We genotyped INDEL, SSR and SNP markers in a sample of 297 rice landraces collected in northern Laos. These glutinous varieties were confirmed to share a loss-of-function mutation in Granule bound starch synthase I (Wx). INDEL markers revealed a high frequency of recombinant genotypes between indica and japonica. Principal component analysis using SSR genotypes of Wx flanking region revealed that glutinous indica landraces were scattered between non-glutinous indica and glutinous-japonica types. High ratios of heterozygosity were found especially in glutinous indica. Haplotype analysis using SNP markers around Wx locus revealed that glutinous indica landraces would have a few chromosome segments of glutinous japonica. Frequent recombinations were confirmed outside of this region in glutinous indica. This intricate genetic structure of landraces suggested that glutinous indica landraces in Laos were generated through repeated natural crossing with glutinous-japonica landraces and severe selection by local farmers. [ABSTRACT FROM AUTHOR]

Hybrid rice has contributed substantially to the improvement of grain production worldwide, yet its poor cooking and tasting characteristics have long been recognized. In this study, 132 recombinant inbred lines derived from LYPJ were used to identify quantitative trait loci ( QTLs) for 12 cooking traits with the high-density SNP linkage map recently developed by our team. We identified 17 QTLs on chromosomes 1, 2, 4, 5, 6, 7, 8, 9 and 11, which accounted for 7.50% to 23.50% of the phenotypic variations. A novel major QTL qBGL7 for boiled grain length was further fine-mapped to an interval of 440 Kb between the two markers RM21906 and gl3 using a BC3F2 population. Two near-isogenic lines with extreme boiled grain length, GX5-176 and GX5-101, could be directly used in improving cooking quality. We also identified a QTL for soaked grain width expansion rate, qSGWE6, in the Wx gene region on chromosome 6. The Wx differential regulation coincided with sequential variation between the two parents. Our work offered a theoretical basis for molecular breeding of high-quality hybrid rice. [ABSTRACT FROM AUTHOR]

The Wx gene primarily influences the physiochemical properties changes of rice (Oryza sativa L.) To improve the grain qualities of the cv. Y58S with low amylose content (AC) value, the genetic effect of Wx gene was explored on quality traits. With the BC3F2 genetic population of Y58S line associated with different AC value as the materials, this study reported the genetic effect of three different Wx alleles (Wxa, Wxin, Wxb) under near-isogenic background. It was shown that Wx had major effects on the quality traits, and the genetic effect value was in the order Wxa > Wxin > Wxb. In addition, during the course of physiochemical properties changes of higher AC rice cultivars determined by Wxa or Wxin allele, accompanied by the increasing of AC, gel consistency and head milled rice recovery would be decreased in combination with the rising of chalky traits, all of which resulted in substantial fluctuations on quality traits (p < 0.01). However, the genetic law was not evident for Wxb allele. Therefore, except for the major gene Wx, the minor genes in the regulative network of starchsynthesis might be utilized for quality improvement. [ABSTRACT FROM AUTHOR]

Wx is considered to be the most important gene controlling eating and cooking qualities and pasting properties in rice (Oryza sativa L.). In this study, a recombinant inbred line population derived from indica rice parents differing in apparent amylose content (AAC) was used to detect quantitative trait loci (QTLs) for ten grain quality parameters for rice quality improvement. QTL mapping was performed on the whole population and on two sub-populations based on Wx genotypes. A total of 29 QTLs were found in the whole population. Ten QTLs for 7 traits were detected in the two sub-populations, four of which (qPRO3.1, qPV9, qHPV9, and qCS7) were also detected in the whole population, whereas the other six were QTLs with minor effects that might be covered by the Wx locus. Besides the Wx locus with the largest effect on AAC and most pasting properties, there were another six QTL clusters contributing to grain quality located on chromosomes 2, 3, 5, 6 and 9. It was also found that some QTLs for peak viscosity, breakdown and consistency were closely linked to rice grain shape related QTLs on chromosome 3. A QTL cluster on chromosome 9 for peak viscosity, hot paste viscosity and cold paste viscosity was detectable in the whole population, which was close to the isoamylase 3 (ISA3) locus. A QTL cluster for both peak time and pasting temperature on chromosome 6 was near to the starch synthase I locus, and was potentially a new QTL with minor effect for peak time and pasting temperature. These findings will promote better understanding of the genetic regulation of rice eating and cooking qualities. [ABSTRACT FROM AUTHOR]

The Wx gene encoding granule-bound starch synthase is responsible for the biosynthesis of amylose, which plays an important role in determining rice eating and cooking quality. However, the functional evaluation of different Wx alleles remains unclear. In the present study, a set of near-isogenic lines (NILs) carrying four different Wx alleles- Wx , Wx , Wx, and wx-was developed in the genetic background of indica rice cv. Yangfunuo 4 (YFN 4), and designated NIL- Wx, NIL- Wx, NIL- Wx, and NIL- wx, respectively. These NILs show great variations in amylose content (AC), gel consistency (GC), starch viscosity, and thermodynamic property. AC showed the following trend among different NILs: NIL- Wx > NIL- Wx > NIL- Wx > NIL- wx, but the opposite trend for GC. NIL- Wx and NIL- Wx had lower starch thermodynamic properties, such as onset temperature, peak temperature, and conclusion temperature, when compared with both NIL- Wx and NIL- wx. The transcript and protein of Wx gene were not detectable in the NIL- wx line. The accumulation levels of Wx transcript and Wx protein were higher in NIL- Wx than those in NIL- Wx and NIL- Wx. Therefore, Wx gene was closely related to starch quality parameters, such as AC, GC, starch paste viscosity, and starch thermodynamic property. Our results provide a solid foundation for rice grain quality improvement through marker-assistant selection. [ABSTRACT FROM AUTHOR]

This study examined the effects of the combinations of Waxy (Wx) and starch branching enzyme IIb (BEIIb) alleles on starch fine structure and functional properties in indica rice cultivars. The results showed that be2b mutant starches with BEIIb deficiency had higher amylose content, shorter amylose long chains, a higher proportion of amylopectin long chains and molecular order, but a lower proportion of amylopectin short chains and relative crystallinity, resulting in higher gelatinization temperature but lower enthalpy and paste viscosity. Compared with the rice lines carrying Wx b allele, Wx a allele contributed to relatively higher amylose content, longer amylopectin chains, less short-range ordered structure and lower relative crystallinity, leading to a little lower gelatinization enthalpy. This study provides new insight into structure-function relations among rice lines with different allele combinations of starch synthesis related genes, which is a useful strategy for rice quality breeding. [Display omitted] [ABSTRACT FROM AUTHOR]

The diversity of the physical and consequently the functional behaviour of starches, isolated from different rice varieties, is related to their specific structures. The latter are directly related to the starch biosynthetic pathway. To fully take advantage of the different functionalities of starches from different rice varieties and to design tailor-made starches, it is important to gain insight into biosynthesis–structure–physical behaviour–functionality relations. In a first part of this review, starch composition is described with a focus on rice starch. Secondly, current knowledge on starch biosynthesis is discussed. This more specifically includes (i) the function of the rice biosynthetic enzymes (i.e. adenosine diphosphate glucose pyrophosphorylases, synthases, branching and debranching enzymes), (ii) the effect of mutations on rice starch structure and, (iii) models for amylose and amylopectin synthesis. Thirdly, starch structure [i.e. from granule (2–100 μm), to growth ring (120–500 nm), blocklet (20–500 nm), amorphous and crystalline lamellae (9 nm), and amylopectin and amylose chain levels (0.1–1.0 nm)] is dealt with. Finally, relations between rice starch structural aspects [i.e. amylopectin (core) chain length distributions] and physical behaviour (i.e. gelatinisation and amylopectin retrogradation as measured by differential scanning calorimetry) are studied. [Copyright &y& Elsevier]

Alignment of the mature protein sequences of the three isoforms of granule-bound starch synthase 1 (GBSS1) of hexaploid wheat Triticum aestivum was used to identify regions showing sequence divergence. Two synthetic peptides based on the Wx-B1 sequence and differing from the corresponding Wx-A1 and Wx-D1 sequences by two or three amino acids were used to immunise rabbits. One peptide elicited an antibody response that was highly selective for the Wx-B1 isoform when used in an indirect enzyme-linked immunosorbent assay (ELISA) format to assay starch granule protein (SGP) preparations prepared from a series of GBSS1 double null lines. A monoclonal antibody specific for GBSS, but not for any one isoform of the enzyme, was subsequently used as the capture antibody in a sandwich ELISA format, with the Wx-B1-selective rabbit polyclonal serum as the detection antibody. The sandwich ELISA format enabled good resolution of Wx-B1b (protein absent) from Wx-B1a (protein present) varieties using direct extraction of antigen from wholemeal flour. A simple method for the solubilisation of SGP using 8 M urea at room temperature was developed. This assay represents a simple, high-throughput means for the identification of wheat lines carrying the Wx-B1b allele that is suitable for testing small samples of starch, flour or crushed grain with a turn-around time of 1 day. [Copyright &y& Elsevier]

PCR-RF-SSCP (PRS), which combines cleaved amplified polymorphic sequence (CAPS) and single-strand conformation polymorphism (SSCP), is expected to be a useful technique for DNA polymorphism analysis. We evaluated the ability of PRS to detect single nucleotide polymorphism (SNP) using the Waxy gene, Wx, of rice, and subsequently were able to identify point mutations in wx mutant lines. The approximately 6-kb Wx gene was divided into five regions for PCR amplification. Two regions, in which most of the point mutations of the wx mutants have been identified, were amplified by PCR and cloned into a vector, and those clones containing SNPs produced as a result of the inherent inaccuracy of PCR were used for the evaluation of PRS. The efficiency of PRS in the detection of SNPs of these clones was over 70%. PRS analysis of the wx genes in 18 waxy mutants was carried out in the five regions using two different restriction endonucleases and two gel conditions, with and without glycerol. Of the 18 lines tested, 17 showed band patterns different from that of the wild type. Most of the mutations identified in this study were nucleotide changes in exons, which result in amino acid changes. One mutation generated an in-frame stop codon, and another was a frame shift mutation by one-base deletion. Two mutations found at a splice site were considered to inhibit normal splicing of mRNA. These results show that PRS is a useful technique for detecting point mutations in large plant genes. [ABSTRACT FROM AUTHOR]

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...TCC(TC)11...(TC)5C(ACC)11, the allele class SSS-B = (AC)3...TCT(TC)6...(TC)4C(ACC)9, and the allele class SSS-C = (AC)3...TCT(TC)6...(TC)4C(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. [ABSTRACT FROM AUTHOR]