scholarly journals Fine‐tuning the amylose content of rice by precise base editing of the Wx gene

2020 ◽  
Vol 19 (1) ◽  
pp. 11-13 ◽  
Author(s):  
Yang Xu ◽  
Qiupeng Lin ◽  
Xiufeng Li ◽  
Fangquan Wang ◽  
Zhihui Chen ◽  
...  
Keyword(s):  
Amylose Content ◽  
Fine Tuning ◽  
Base Editing ◽  
Wx Gene

UTILIZATION OF DNA MARKERS FOR EVALUATING AMYLOSE CONTENT OF GRAIN IN RICE

2013 ◽  
Vol 83 (5) ◽  
Author(s):  
Dương Thanh Thủy ◽  
Taiichiro Ookawa
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Amylose Content ◽  
Waxy Gene ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Branching Enzymes ◽  
Wx Gene ◽  
High Amylose ◽  
Suitable Marker ◽  
Granule Bound Starch Synthase

The sensory and functional properties of rice are predominantly associated with its amylose content. Granule-bound starch synthase (GBSS) encoded by the Waxy (Wx) gene determines the synthesis of amylose, while starch branching enzymes encoded by Sbe genes are involved in the formation of amylopectin. Some studies have demonstrated that Wx gene is the major controller of amylose content but there are one or more modifying genes affecting the amylose content. Three markers,  microsatellite, Single – nucleotide – polymorphism (G/T SNP) in Wx gene and Single – nucleotide – polymorphism (T/C SNP) in Sbe1 gene, were tested for their association with amylose content using sixty-nine  rice accessions from twenty countries. Of the three markers, two markers in Wx gene are significantly associated with amylose content. The combination of two markers in Wx gene (haplotypes) explained 83.8% of the variation in amylose content and discriminated the three market classes of glutinous, low, intermediate and high amylose content of rice from each other. And T/C SNP in Sbe1 locus was not a suitable marker for amylose content. Keywords: marker, amylose content, Waxy gene.

scholarly journals Wxlv, the Ancestral Allele of Rice Waxy Gene

Proceedings ◽  
2020 ◽  
Vol 36 (1) ◽  
pp. 140 ◽  
Author(s):  
Changquan Zhang ◽  
Jihui Zhu ◽  
Shengjie Chen ◽  
Qiaoquan Liu
Keyword(s):  
Amylose Content ◽  
Waxy Gene ◽  
Evolutionary Analysis ◽  
Ancestral Allele ◽  
Cultivated Rice ◽  
Functional Sites ◽  
Wx Locus ◽  
Wx Gene ◽  
Shed Light

In rice endosperms, the Waxy (Wx) gene is important for amylose synthesis, and various Wx alleles control the amylose content and affect the taste of cooked rice. Herein, we report the cloning of the ancestral allele Wxlv of the Wx locus, which affects the mouthfeel of rice grains by modulating the size of amylose molecules. Using evolutionary analysis, we demonstrated that Wxlv originated directly from wild rice, and the three major Wx alleles in cultivated rice (Wxb, Wxa, and Wxin) differentiated after the substitution of one base pair at the functional sites. These data indicate that the Wxlv allele played an important role in artificial selection and domestication. The findings also shed light on the evolution of various Wx alleles, which have greatly contributed to improving the eating and cooking quality of rice.

scholarly journals Analysis of the relationship between Wx gene polymorphisms and amylose content in hulless barley  

2017 ◽  
Vol 53 (No. 4) ◽  
pp. 144-152 ◽  
Author(s):  
K. Wu ◽  
X. Yao ◽  
Y. Yao ◽  
D. Chi ◽  
Z. Feng
Keyword(s):  
Gene Polymorphisms ◽  
Genomic Dna ◽  
Gene Locus ◽  
Amylose Content ◽  
Product Size ◽  
Hulless Barley ◽  
Pcr Product ◽  
Wx Gene ◽  
The Relationship ◽  
Selection Of

I<sub>2</sub>-KI staining was used to phenotype 151 hulless barley plants, which determined that five samples were of the waxy variety, namely 14-Z152, IG107028, Puebla, Hu Zhu Shuang Cao Ren, and APM-HC1905. Using the dual-wavelength method, the average amylose content of the 151 samples was 25.9%, ranging from 4.9 to 38.5%. The average amylose content of the five waxy varieties was 14.3%, ranging from 4.9 to 18.6%. Genomic DNA from 48 samples showing a significantly variable amylose content was used as template and PCR amplified using primer pair P4. Statistical analysis indicated that the PCR product size positively correlated with amylose content. The Wx gene locus was determined to be polymorphic, and was positively correlated with amylose. Based on the electrophoresis results, the 48 samples were divided into 4 types. PCR product types I, II, III, and IV were 457, 481, 489, and 491 bp in length, respectively, with the respective amylose content ranges of 4.9–27%, 29–30%, 31–35%, and 36–38%. Primer P4 can be used as a complementary marker for the selection of different amylose content hulless barley germplasms.

scholarly journals CRISPR-Based Genome Editing Tools: Insights into Technological Breakthroughs and Future Challenges

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 797
Author(s):  
Muntazir Mushtaq ◽  
Aejaz Ahmad Dar ◽  
Milan Skalicky ◽  
Anshika Tyagi ◽  
Nancy Bhagat ◽  
...  
Keyword(s):  
Genome Editing ◽  
Crop Improvement ◽  
Fine Tuning ◽  
Plant Genome ◽  
Base Editing ◽  
Global Food Security ◽  
Technological Advances ◽  
Efficient Delivery ◽  
Future Challenges ◽  
Recent Developments

Genome-editing (GE) is having a tremendous influence around the globe in the life science community. Among its versatile uses, the desired modifications of genes, and more importantly the transgene (DNA)-free approach to develop genetically modified organism (GMO), are of special interest. The recent and rapid developments in genome-editing technology have given rise to hopes to achieve global food security in a sustainable manner. We here discuss recent developments in CRISPR-based genome-editing tools for crop improvement concerning adaptation, opportunities, and challenges. Some of the notable advances highlighted here include the development of transgene (DNA)-free genome plants, the availability of compatible nucleases, and the development of safe and effective CRISPR delivery vehicles for plant genome editing, multi-gene targeting and complex genome editing, base editing and prime editing to achieve more complex genetic engineering. Additionally, new avenues that facilitate fine-tuning plant gene regulation have also been addressed. In spite of the tremendous potential of CRISPR and other gene editing tools, major challenges remain. Some of the challenges are related to the practical advances required for the efficient delivery of CRISPR reagents and for precision genome editing, while others come from government policies and public acceptance. This review will therefore be helpful to gain insights into technological advances, its applications, and future challenges for crop improvement.

scholarly journals High-Efficiency Reduction of Rice Amylose Content via CRISPR/Cas9-Mediated Base Editing

Rice Science ◽  
2020 ◽  
Vol 27 (6) ◽  
pp. 445-448
Author(s):  
Li He ◽  
Li Xiufeng ◽  
Xu Yang ◽  
Liu Hualong ◽  
He Mingliang ◽  
...  
Keyword(s):  
High Efficiency ◽  
Amylose Content ◽  
Base Editing

scholarly journals In-situ generation of large numbers of genetic combinations for metabolic reprogramming via CRISPR-guided base editing

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yu Wang ◽  
Haijiao Cheng ◽  
Yang Liu ◽  
Ye Liu ◽  
Xiao Wen ◽  
...  
Keyword(s):  
Large Scale ◽  
High Efficiency ◽  
Metabolic Reprogramming ◽  
Fine Tuning ◽  
Ex Situ ◽  
Library Size ◽  
Base Editing ◽  
Large Numbers ◽  
Multigene Expression

AbstractReprogramming complex cellular metabolism requires simultaneous regulation of multigene expression. Ex-situ cloning-based methods are commonly used, but the target gene number and combinatorial library size are severely limited by cloning and transformation efficiencies. In-situ methods such as multiplex automated genome engineering (MAGE) depends on high-efficiency transformation and incorporation of heterologous DNA donors, which are limited to few microorganisms. Here, we describe a Base Editor-Targeted and Template-free Expression Regulation (BETTER) method for simultaneously diversifying multigene expression. BETTER repurposes CRISPR-guided base editors and in-situ generates large numbers of genetic combinations of diverse ribosome binding sites, 5’ untranslated regions, or promoters, without library construction, transformation, and incorporation of DNA donors. We apply BETTER to simultaneously regulate expression of up to ten genes in industrial and model microorganisms Corynebacterium glutamicum and Bacillus subtilis. Variants with improved xylose catabolism, glycerol catabolism, or lycopene biosynthesis are respectively obtained. This technology will be useful for large-scale fine-tuning of multigene expression in both genetically tractable and intractable microorganisms.

scholarly journals Generation of a New Glutinous Photothermosensitive Genic-Male-Sterile (PTGMS) Line by CRISPR/Cas9-Directed Mutagenesis of Wx in Rice (Oryza sativa L.)

Agriculture ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 1044
Author(s):  
Kaichong Teng ◽  
Xin Wang ◽  
Xinying Guo ◽  
Yaoguang Liu ◽  
Rongbai Li
Keyword(s):  
Mutation Rate ◽  
Hybrid Rice ◽  
Amylose Content ◽  
Rice Variety ◽  
High Yield ◽  
Male Sterile ◽  
High Quality ◽  
Glutinous Rice ◽  
Homozygous Mutant ◽  
Wx Gene

The Photothermosensitive Genic-Male-Sterile (PTGMS) line, Y58S, an indica rice variety, combines high-quality and high-light-efficiency use, disease and stress resistance, and excellent plant type and mating force. Y58S is widely used to assemble two-line hybrid rice varieties, especially super hybrids. The Wx gene is the main effector gene for controlling amylose synthesis, which determines the amylose content (AC) of rice grains. By editing this gene, a glutinous line with a low AC can be obtained. In this study, the CRISPR/Cas9 system was used to mediate the editing of the Wx gene, which caused ultra-low AC mutations that produced a PTGMS glutinous rice strain with excellent waxiness. The results showed that 18 positively transformed plants were obtained from the T0 generation, with a mutation rate of 64.29%, of which six were homozygous mutant plants, indicating that the gene-editing target had a higher targeting efficiency and a higher homozygosity mutation rate. Compared to the wild type, the AC of the mutants was significantly lower. Through molecular marker detection and screening of T1 and T2 generations, five homozygous T-DNA-free mutant strains were identified that were consistent with Y58S in fertility and other agronomic traits except for AC. Among these, the AC of the W-1-B-5 homozygous mutant, the glutinous PTGMS line wx-Y58S, was as low as 0.6%. Our research revealed that the Wx gene of excellent PTGMS rice can be edited to generate a new waxy PTGMS line using the CRISPR/Cas9 system. This study provided a simple and effective strategy for breeding high-yield, high-quality, and glutinous two-line hybrid rice, and provided excellent sterile lines for their large-scale application. Once put into use, waxy hybrid rice will greatly improve the yield of glutinous rice and increase social benefits.

scholarly journals Hd1 Allele Types and Their Associations with Major Agronomic Traits in Korean Rice Cultivars

Plants ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2408
Author(s):  
Youngjun Mo ◽  
Chang-Min Lee ◽  
Hyang-Mi Park ◽  
Su-Kyung Ha ◽  
Mi-Jung Kim ◽  
...  
Keyword(s):  
Agronomic Traits ◽  
Amylose Content ◽  
Heading Date ◽  
Fine Tuning ◽  
Rice Cultivars ◽  
Protein Coding ◽  
Coding Regions ◽  
Days To Heading ◽  
Wide Range ◽  
Korean Rice

Optimizing flowering time in crop plants is critical for maximizing yield and quality under target environments. While there is a wide range of heading date variation in Korean rice cultivars, the underlying gene mechanisms are unclear. Here, we sequenced the protein coding regions of Hd1, the major rice heading date gene, from 293 Korean rice cultivars and investigated the associations between Hd1 allele types and major agronomic traits under four different environments. There were four functional Hd1 and five nonfunctional hd1 alleles distributed among the 293 Korean rice cultivars. The effects of the Hd1 allele types were highly significant for days to heading in all four environments, explaining 51.4–65.8% of the phenotypic variation. On average, cultivars carrying nonfunctional hd1 headed 13.7 days earlier than those carrying functional Hd1. While the Hd1 allele types exhibited highly significant effects on culm length and protein content under all four environments, the differences between cultivars carrying Hd1 and hd1 were minimal. The effects of the Hd1 allele types on amylose content were significant in only one of the four environments. Our results provide useful information for fine-tuning rice heading dates by utilizing different Hd1 alleles in rice breeding programs.

scholarly journals Waxy Gene-Orthologs in Wheat × Thinopyrum Amphidiploids

Agronomy ◽  
2020 ◽  
Vol 10 (7) ◽  
pp. 963
Author(s):  
Marina V. Klimushina ◽  
Pavel Yu. Kroupin ◽  
Mikhail S. Bazhenov ◽  
Gennady I. Karlov ◽  
Mikhail G. Divashuk
Keyword(s):  
Amylose Content ◽  
Wide Spectrum ◽  
Allelic Diversity ◽  
Wheat Breeding ◽  
Raw Material ◽  
Waxy Gene ◽  
Allelic Polymorphism ◽  
Wx Gene ◽  
Main Component ◽  
Gene Orthologs

Starch, as the main component of grain in cereals, serves as the major source of calories in staple food and as a raw material for industry. As the technological and digestive properties of starch depend on its content, the management of its components, amylose and amylopectin, is of great importance. The starch properties of wheat grain can be attuned using allelic variations of genes, including granule-bound starch synthase I (GBSS I), or Wx. The tertiary gene pool, including wheatgrass (Thinopyrum) species, provides a wide spectrum of genes-orthologs that can be used to increase the allelic diversity of wheat genes by wide hybridization. Octaploid partial wheat–wheatgrass hybrids (amphidiploids, WWGHs) combine the complete genome of bread wheat (BBAADD), and a mixed genome from the chromosomes of intermediate wheatgrass (Thinopyrum intermedium, genomic composition JrJrJvsJvsStSt) and tall wheatgrass (Th. ponticum, JJJJJJJsJsJsJs). Thus, WWGHs may carry Wx genes not only of wheat (Wx-B1, Wx-A1 and Wx-D1) but also of wheatgrass origin. We aimed to assess the level of amylose in starch and investigate the polymorphism of Wx genes in 12 accessions of WWGHs. Additionally, we characterized orthologous Wx genes in the genomes of wild wheat-related species involved in the development of the studied WWGHs, Th. intermedium and Th. ponticum, as well as in the putative donors of their subgenomes, bessarabian wheatgrass (Th. bessarabicum, JbJb) and bluebunch wheatgrass (Pseudoroegneria stipifolia, St1St1St2St2). Although no significant differences in amylose content were found between different WWGH accessions, SDS-PAGE demonstrated that at least two WWGHs have an additional band. We sequenced the Wx gene-orthologs in Th. bessarabicum, P. stipifolia, Th. intermedium and Th. ponticum, and developed a WXTH marker that can discriminate the Thinopyrum Wx gene in the wheat background, and localized it to the 7E chromosome in Th. elongatum. Using the WXTH marker we revealed the allelic polymorphism of the Thinopyrum Wx gene in the studied WWGHs. The applicability of Thinopyrum Wx genes in wheat breeding and their effect on starch quality are discussed.

CRISPR/Cas Genome Editing and Precision Plant Breeding in Agriculture

2019 ◽  
Vol 70 (1) ◽  
pp. 667-697 ◽  
Author(s):  
Kunling Chen ◽  
Yanpeng Wang ◽  
Rui Zhang ◽  
Huawei Zhang ◽  
Caixia Gao
Keyword(s):  
Plant Breeding ◽  
Genome Editing ◽  
Crop Improvement ◽  
Delivery Systems ◽  
Fine Tuning ◽  
Plant Genome ◽  
Nucleotide Substitutions ◽  
Base Editing ◽  
Homology Directed Repair ◽  
Challenges And Opportunities

Enhanced agricultural production through innovative breeding technology is urgently needed to increase access to nutritious foods worldwide. Recent advances in CRISPR/Cas genome editing enable efficient targeted modification in most crops, thus promising to accelerate crop improvement. Here, we review advances in CRISPR/Cas9 and its variants and examine their applications in plant genome editing and related manipulations. We highlight base-editing tools that enable targeted nucleotide substitutions and describe the various delivery systems, particularly DNA-free methods, that have linked genome editing with crop breeding. We summarize the applications of genome editing for trait improvement, development of techniques for fine-tuning gene regulation, strategies for breeding virus resistance, and the use of high-throughput mutant libraries. We outline future perspectives for genome editing in plant synthetic biology and domestication, advances in delivery systems, editing specificity, homology-directed repair, and gene drives. Finally, we discuss the challenges and opportunities for precision plant breeding and its bright future in agriculture.

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