Search Results (9)

Background: The coordinated improvement of yield, quality and resistance is a primary goal in rice breeding. Gene editing technology is a novel method for precise multiplex gene improvement. Methods: In this study, we constructed a multiplex CRISPR/Cas9 vector targeting yield-related genes (GS3, OsPIL15, Gn1a), fragrance gene (OsBADH2) and rice blast resistance gene (Pi21) to pyramid traits for enhanced yield, quality, and disease resistance in rice. A tRNA-assisted CRISPR/Cas9 multiplex gene editing vector, M601-OsPIL15/GS3/Gn1a/OsBADH2/Pi21-gRNA, was constructed. Genetic transformation was performed using the Agrobacterium-mediated method with the japonica rice variety Xin Dao 53 as the recipient. Mutation editing efficiency was detected in T₀ transgenic plants. Grain length, grain number per panicle, thousand-grain weight, 2-acetyl-1-pyrroline (2-AP) content, and rice blast resistance of homozygous lines were measured in the T₃ generations. Results: Effectively edited plants were obtained in the T₀ generation. The simultaneous editing efficiency for all five genes reached 9.38%. The individual gene editing efficiencies for Pi21, GS3, OsBADH2, Gn1a, and OsPIL15 were 78%, 63%, 56%, 54%, and 13%, respectively. Five five-gene homozygous edited lines with two genotypes were selected in the T₂ generation. In the T₃ generation, compared with the wild-type (WT), the edited homozygous lines showed increased grain number per panicle (14.60–25.61%), increased grain length (7.39–11.16%), increased grain length–width ratio (8.37–13.02%), increased thousand-grain weight (3.79–9.15%), a 42–64 folds increase in the fragrant substance 2-AP content, and significantly enhanced rice blast resistance. Meanwhile, there were no significant changes in other agronomic traits. Conclusions: CRISPR/Cas9-mediated multiplex gene editing technology enabled the simultaneous editing of genes related to rice yield, quality, and disease resistance. This provides an effective approach for obtaining new japonica rice germplasm with blast resistance, long grains, and fragrance. Full article

Fragrant rice is regarded as a premium variety due to its distinctive aroma, delicate texture, and rich nutritional value. This aroma primarily originates from 2-acetyl-1-pyrroline (2-AP), but the metabolic basis of 2-AP remains elusive to this day, and the genetic basis for metabolite accumulation is largely unknown. While several researchers have investigated differences in 2-AP synthesis pathways between fragrant and non-fragrant rice, few studies have examined the 2-AP synthesis pathways in fragrant rice varieties exhibiting 2-AP differences. Therefore, after conducting gene similarity analyses on six fragrant rice varieties, we measured the expression levels of substances and related genes involved in multiple metabolic pathways within the 2-AP synthesis pathway, along with the specific enzyme activities associated with these pathways. Results indicate that XG12 (Guizhou Fragrant Rice Variety) exhibits the highest 2-AP content, yet its efficiency in synthesizing 2-AP is not the highest across any individual metabolic pathway. This finding reveals that among fragrant rice varieties, 2-AP content negatively correlates with OsBadh2 expression levels and GABA content, while showing no linear correlation with other related substances or metabolic genes. At this point, variations in the major effect gene OsBadh2 no longer dominate; instead, subtle differences in 2-AP content are jointly determined by numerous minor effect genes and environmental factors. This phenomenon not only resolves apparent contradictions but profoundly illuminates the complex regulatory mechanisms governing 2-AP biosynthesis. 2-AP synthesis represents a dynamic equilibrium process, with different fragrant rice varieties potentially accumulating 2-AP through distinct metabolic pathways. Additionally, this study analyzed the volatile organic compounds (VOCs) of six fragrant rice varieties through metabolomics. Results revealed that DLX, which exhibited the lowest 2-AP content, contained the richest array of aggregated VOCs, indicating no correlation between 2-AP and numerous VOCs. Our findings provide a clear research direction for elucidating the genetic regulatory mechanisms of 2-AP underlying fragrant rice and lay the foundation for technological research aimed at enhancing the aroma of fragrant rice varieties. Full article

In addition to being governed by genetic factors, environmental factors also play a crucial role in influencing the fragrance of rice. In this research, the high-quality rice variety Dalixiang was selected as the experimental material to investigate the impacts of soil nutrients in Guiyang and Meitan on its fragrance. The results indicated that the levels of ammonium nitrogen, organic matter, total nitrogen, available nitrogen, and the pH value in the soil of Meitan were lower compared to those in Guiyang. Conversely, the contents of total potassium, available phosphorus, and available potassium were higher in Meitan. Specifically, the concentrations of 2-acetyl-1-pyrroline (2AP) in the leaves of Dalixiang at the heading stage and in the grains at the maturity stage at the Meitan planting site were 0.13 mg/kg and 0.56 mg/kg, respectively. These values were significantly lower than the 0.17 mg/kg and 0.64 mg/kg measured at the Guiyang planting site. This phenomenon is associated with the higher expression levels of the betaine aldehyde dehydrogenase (OsBadh2) gene, enhanced enzyme activities, and a greater content of γ-aminobutyric acid (GABA) in the leaves of Dalixiang at the Meitan planting site. In contrast, the expression levels of genes related to triose phosphate isomerase (OsTPI), proline dehydrogenase (OsProDH), ornithine aminotransferase (OsOAT), and Delta1-pyrroline-5-carboxylic acid synthetase (OsP5CS), along with their corresponding enzyme activities, as well as the contents of methylglyoxal, proline, and ornithine, were lower. In conclusion, due to the influence of the Guiyang environment, the biosynthesis of Dalixiang 2AP was promoted, which made the Dalixiang planted in Guiyang stronger than that planted in Meitan. This study provides a theoretical basis for the selection of the best planting area of Dalixiang and the improvement of Dalixiang flavor through agronomic cultivation techniques. Full article

‘Dianheyou615’ (DHY615) is an elite Dian (D1)-type hybrid japonica rice variety, renowned for its high yield, exceptional grain quality, and unique adaptability to both irrigated and rainfed conditions in the Yungui Plateau of southwestern China. However, the genetic mechanisms underlying the agronomic performance of the D1-type hybrid japonica rice remain unclear. In this study, a comprehensive analysis of ‘DHY615’’s agronomic performance, genetic genealogy, and molecular genetic foundation was conducted to dissect its desirable traits for upland rainfed cultivation across diverse ecological environments. The main findings indicate that ‘DHY615’ possesses 6432 heterozygous SNPs, with 57.48% and 14.43% located in the promoter and coding regions, respectively, potentially affecting key phenotypic traits. High-impact SNPs variants and numerous well-known functional genes were identified, such as OsAAP6, GS3, Sd1, Rf1, BADH2, BPh14, Rymv1, OsFRO1, NRT1.1B, SKC1, OsNCED2, and qUVR-10, which are likely linked to traits including plant architecture, grain yield, grain quality, and resistance to various biotic and abiotic stresses (e.g., disease, cold, drought, salt, high iron, and high UV radiation). Notably, ‘Nan615’ harbors a greater number of functional allele variants compared to ‘H479A’, which potentially explaining its superior grain yield and remarkable adaptability. This study offers novel and valuable insights into the molecular genetic foundation of the plateau D1-type hybrid japonica rice, underscoring its potential for sustainable rice production across diverse ecological zones, especially with its unparalleled high-altitude adaptability to rainfed upland planting. Full article

Basmati rice is a premium aromatic rice that consumers choose primarily because of its distinct aroma and excellent grain quality. The grain quality of Basmati rice (GQBR) reflects the perspectives of producers, processors, sellers, and consumers related to the production, processing, marketing, and consumption of Basmati rice. Consumers, an invaluable part of the production demand and value chain of the Basmati rice industry, have the freedom to choose from different types of aromatic rice. Consumers expect their preferred Basmati rice to possess all superior rice grain qualities, including the physical, biochemical, and physiological properties. Gene functional analysis explained that a 10-base pair deletion in the promoter region of the OsSPL16 gene causes the slender grains in Basmati rice, whereas an 8-base-pair deletion in exon 7 of the OsBadh2 gene (located in the fgr region on rice chromosome 8) results in the distinct aroma. Furthermore, a combination of the genetic characteristics of the gw8 and gs3 genes has led to the creation of a long-grain Basmati-type rice cultivar. It has also been demonstrated that agricultural, genetic, and environmental conditions significantly influence GQBR. Hence, research on improving GQBR requires a multidimensional approach and sophisticated elements due to the complexity of its nature and preference diversity. This review covers the basic definitions of grain quality traits, consumer preference criteria, influencing factors, and strategies for producing superior-quality Basmati rice in the United States. This knowledge will be useful in improving the grain quality of Basmati and Basmati-type rice, as well as developing appropriate breeding programs that will meet the preferences of different countries and cultures. Full article

The most important volatile in determining the aroma of fragrant rice is 2-Acetyl-1-pyrroline (2-AP); however, the transcriptional regulation mechanism of 2-AP biosynthesis in fragrant rice is still unclear. In this study, Osp5cs1 knockout mutant lines and OsP5CS1 over-expression lines were constructed by the genetic transformation of the Indica rice cultivar, i.e., ‘Zhonghua11′, which knocks out OsBADH2 to produce fragrance in aromatic rice. The OsP5CS1 gene was also identified as a key gene in the 2-AP biosynthesis pathway of aromatic rice. The OsP5CS1 promoter was used as bait, and the OsbZIP60-like transcription factor was screened by yeast one-hybrid assays. The OsbZIP60-like transcription factor specifically bound to the OsP5CS1 gene. The dual luciferase reporting system found that the OsbZIP60-like transcription factor promoted the transcriptional activation of OsP5CS1. Compared with the wild type, OsP5CS1 gene expression was significantly down-regulated in the Osbzip60-like mutant and resulted in a substantial reduction in 2-AP biosynthesis. Moreover, the OsP5CS1 gene expression was significantly up-regulated in OsbZIP60-like over-expressed plants, and the 2-AP concentrations were also increased, whereas the Osbzip60-like mutants were found to be sensitive to Zn deficiency. Overall, the OsbZIP60-like transcription factor promoted the 2-AP accumulation. This study provides a theoretical basis for the transcriptional regulation mechanism of 2-AP biosynthesis and explores the function of the OsbZIP transcription factor in fragrant rice. Full article

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. Full article

Mutations in the Betaine aldehyde dehydrogenase 2 (OsBadh2) gene resulted in aroma, which is a highly preferred grain quality attribute in rice. However, research on naturally occurring aromatic rice has revealed ambiguity and controversy regarding aroma emission, stress tolerance, and response to salinity. In this study, mutant lines of two non-aromatic varieties, Huaidao#5 (WT_HD) and Jiahua#1 (WT_JH), were generated by targeted mutagenesis of OsBadh2 using CRISPR/Cas9 technology. The mutant lines of both varieties became aromatic; however, WT_HD mutants exhibited an improved tolerance, while those of WT_JH showed a reduced tolerance to salt stress. To gain insight into the molecular mechanism leading to the opposite effects, comparative analyses of the physiological activities and expressions of aroma- and salinity-related genes were investigated. The WT_HD mutants had a lower mean increment rate of malondialdehyde, superoxide dismutase, glutamate, and proline content, with a higher mean increment rate of γ-aminobutyric acid, hydrogen peroxide, and catalase than the WT_JH mutants. Fluctuations were also detected in the salinity-related gene expression. Thus, the response mechanism of OsBadh2 mutants is complicated where the genetic makeup of the rice variety and interactions of several genes are involved, which requires more in-depth research to explore the possibility of producing highly tolerant aromatic rice genotypes. Full article

The significant increase in grain yield and quality are often antagonistic but a constant demand for breeders and consumers. Some genes related to cytochrome P450 family are known for rice organ growth but their role in controlling grain yield is still unknown. Here, we generated new rice mutants with high yield and improved aroma by simultaneously editing three cytochrome P450 homoeologs (Os03g0603100, Os03g0568400, and GL3.2) and OsBADH2 with the CRISPR/Cas9 system, and RNA-sequencing and proteomic analysis were performed to unveil the subsequent changes. High mutation efficiency was achieved in both target sites of each gene and the mutations were predominantly only deletions, while insertions were rare, and no mutations were detected in the five most likely off-target sites against each sgRNA. Mutants exhibited increased grain size, 2-acetyl-1-pyrroline (2AP) content, and grain cell numbers while there was no change in other agronomic traits. Transgene-DNA-free mutant lines appeared with a frequency of 44.44% and homozygous mutations were stably transmitted, and bi-allelic and heterozygous mutations followed Mendelian inheritance, while the inheritance of chimeric mutations was unpredictable. Deep RNA sequencing and proteomic results revealed the regulation of genes and proteins related to cytochrome P450 family, grain size and development, and cell cycle. The KEGG and hub-gene and protein network analysis showed that the gene and proteins related to ribosomal and photosynthesis pathways were mainly enriched, respectively. Our findings provide a broad and detailed basis to understand the role of CRISPR/Cas9 in rice yield and quality improvement. Full article