Polyamines (PAs) are pleiotropic bioorganic molecules. Cellular PA contents are determined by a balance between PA synthesis and degradation. PAs have been extensively demonstrated to play vital roles in the modulation of plant developmental processes and adaptation to various environmental stresses. In this review, the latest advances on the diverse roles of PAs in a range of developmental processes, such as morphogenesis, organogenesis, growth and development, and fruit ripening, are summarized and discussed. Besides, the crosstalk between PAs and phytohormones or other signalling molecules, including H2O2 and NO, involved in these processes is dwelled on. In addition, the attempts made to improve the yield and quality of grain and vegetable crops through altering the PA catabolism are enumerated. Finally, several other vital questions that remain unanswered are proposed and discussed. These include the mechanisms underlying the cooperative regulation of developmental processes by PAs and their interplaying partners like phytohormones, H2O2 and NO; PA transport for maintaining homeostasis; and utilization of PA anabolism/catabolism for generating high-yield and good-quality crops. This review aims to gain new insights into the pleiotropic role of PAs in the modulation of plant growth and development, which provides an alternative approach for manipulating and engineering valuable crop varieties that can be used in the future.
多胺（PA）是多效生物有机分子。细胞中的多胺含量取决于多胺合成和降解之间的平衡。大量研究表明，多胺在调节植物发育过程和适应各种环境胁迫方面发挥着重要作用。本综述总结并讨论了 PAs 在形态发生、器官发生、生长发育和果实成熟等一系列发育过程中的不同作用的最新进展。此外，还详细介绍了 PAs 与植物激素或其他信号分子（包括 H2O2 和 NO）在这些过程中的相互影响。此外，还列举了通过改变 PA 分解代谢来提高谷物和蔬菜作物产量和质量的尝试。最后，还提出并讨论了其他几个尚未解答的重要问题。这些问题包括 PA 及其相互作用伙伴（如植物激素、H2O2 和 NO）对发育过程的协同调控机制；维持平衡的 PA 运输；以及利用 PA 合成代谢/代谢产生高产优质作物。本综述旨在对 PA 在调节植物生长和发育过程中的多效性作用获得新的认识，从而为操纵和改造未来可利用的有价值作物品种提供另一种方法。
{"title":"Polyamines: pleiotropic molecules regulating plant development and enhancing crop yield and quality","authors":"Haishan Yang, Yinyin Fang, Zhiman Liang, Tian Qin, Ji-Hong Liu, Taibo Liu","doi":"10.1111/pbi.14440","DOIUrl":"10.1111/pbi.14440","url":null,"abstract":"<p>Polyamines (PAs) are pleiotropic bioorganic molecules. Cellular PA contents are determined by a balance between PA synthesis and degradation. PAs have been extensively demonstrated to play vital roles in the modulation of plant developmental processes and adaptation to various environmental stresses. In this review, the latest advances on the diverse roles of PAs in a range of developmental processes, such as morphogenesis, organogenesis, growth and development, and fruit ripening, are summarized and discussed. Besides, the crosstalk between PAs and phytohormones or other signalling molecules, including H<sub>2</sub>O<sub>2</sub> and NO, involved in these processes is dwelled on. In addition, the attempts made to improve the yield and quality of grain and vegetable crops through altering the PA catabolism are enumerated. Finally, several other vital questions that remain unanswered are proposed and discussed. These include the mechanisms underlying the cooperative regulation of developmental processes by PAs and their interplaying partners like phytohormones, H<sub>2</sub>O<sub>2</sub> and NO; PA transport for maintaining homeostasis; and utilization of PA anabolism/catabolism for generating high-yield and good-quality crops. This review aims to gain new insights into the pleiotropic role of PAs in the modulation of plant growth and development, which provides an alternative approach for manipulating and engineering valuable crop varieties that can be used in the future.</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3194-3201"},"PeriodicalIF":10.1,"publicationDate":"2024-07-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14440","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141722760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Yazhou Yang, Jia Liu, Stacy D. Singer, Guohua Yan, Dennis R. Bennet, Yue Liu, Jean-Michel Hily, Weirong Xu, Yingzhen Yang, Xiping Wang, Gan-Yuan Zhong, Zhongchi Liu, Yong-Qiang Charles An, Huawei Liu, Zongrang Liu
Cis-regulatory elements (CREs) are integral to the spatiotemporal and quantitative expression dynamics of target genes, thus directly influencing phenotypic variation and evolution. However, many of these CREs become highly susceptible to transcriptional silencing when in a transgenic state, particularly when organised as tandem repeats. We investigated the mechanism of this phenomenon and found that three of the six selected flower-specific CREs were prone to transcriptional silencing when in a transgenic context. We determined that this silencing was caused by the ectopic expression of non-coding RNAs (ncRNAs), which were processed into 24-nt small interfering RNAs (siRNAs) that drove RNA-directed DNA methylation (RdDM). Detailed analyses revealed that aberrant ncRNA transcription within the AGAMOUS enhancer (AGe) in a transgenic context was significantly enhanced by an adjacent CaMV35S enhancer (35Se). This particular enhancer is known to mis-activate the regulatory activities of various CREs, including the AGe. Furthermore, an insertion of 35Se approximately 3.5 kb upstream of the AGe in its genomic locus also resulted in the ectopic induction of ncRNA/siRNA production and de novo methylation specifically in the AGe, but not other regions, as well as the production of mutant flowers. This confirmed that interactions between the 35Se and AGe can induce RdDM activity in both genomic and transgenic states. These findings highlight a novel epigenetic role for CRE–CRE interactions in plants, shedding light on the underlying forces driving hypermethylation in transgenes, duplicate genes/enhancers, and repetitive transposons, in which interactions between CREs are inevitable.
{"title":"Ectopic enhancer–enhancer interactions as causal forces driving RNA-directed DNA methylation in gene regulatory regions","authors":"Yazhou Yang, Jia Liu, Stacy D. Singer, Guohua Yan, Dennis R. Bennet, Yue Liu, Jean-Michel Hily, Weirong Xu, Yingzhen Yang, Xiping Wang, Gan-Yuan Zhong, Zhongchi Liu, Yong-Qiang Charles An, Huawei Liu, Zongrang Liu","doi":"10.1111/pbi.14435","DOIUrl":"10.1111/pbi.14435","url":null,"abstract":"<p><i>Cis</i>-regulatory elements (<i>CREs</i>) are integral to the spatiotemporal and quantitative expression dynamics of target genes, thus directly influencing phenotypic variation and evolution. However, many of these <i>CREs</i> become highly susceptible to transcriptional silencing when in a transgenic state, particularly when organised as tandem repeats. We investigated the mechanism of this phenomenon and found that three of the six selected flower-specific <i>CREs</i> were prone to transcriptional silencing when in a transgenic context. We determined that this silencing was caused by the ectopic expression of non-coding RNAs (ncRNAs), which were processed into 24-nt small interfering RNAs (siRNAs) that drove RNA-directed DNA methylation (RdDM). Detailed analyses revealed that aberrant ncRNA transcription within the <i>AGAMOUS</i> enhancer (<i>AGe</i>) in a transgenic context was significantly enhanced by an adjacent <i>CaMV35S</i> enhancer (<i>35Se</i>). This particular enhancer is known to mis-activate the regulatory activities of various <i>CREs,</i> including the <i>AGe</i>. Furthermore, an insertion of <i>35Se</i> approximately 3.5 kb upstream of the <i>AGe</i> in its genomic locus also resulted in the ectopic induction of ncRNA/siRNA production and <i>de novo</i> methylation specifically in the <i>AGe</i>, but not other regions, as well as the production of mutant flowers. This confirmed that interactions between the <i>35Se</i> and <i>AGe</i> can induce RdDM activity in both genomic and transgenic states. These findings highlight a novel epigenetic role for <i>CRE–CRE</i> interactions in plants, shedding light on the underlying forces driving hypermethylation in transgenes, duplicate genes/enhancers, and repetitive transposons, in which interactions between <i>CREs</i> are inevitable.</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3121-3134"},"PeriodicalIF":10.1,"publicationDate":"2024-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14435","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141632125","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Jun-Hye Shin, Sera Oh, Mi-Hwa Jang, Seok-Yong Lee, Chanhong Min, Young-Jae Eu, Hilal Begum, Jong-Chan Kim, Gap Ryol Lee, Han-Bin Oh, Matthew J. Paul, Julian K.-C. Ma, Ho-Shin Gwak, Hyewon Youn, Seong-Ryong Kim
For several decades, a plant-based expression system has been proposed as an alternative platform for the production of biopharmaceuticals including therapeutic monoclonal antibodies (mAbs), but the immunogenicity concerns associated with plant-specific N-glycans attached in plant-based biopharmaceuticals has not been completely solved. To eliminate all plant-specific N-glycan structure, eight genes involved in plant-specific N-glycosylation were mutated in rice (Oryza sativa) using the CRISPR/Cas9 system. The glycoengineered cell lines, PhytoRice®, contained a predominant GnGn (G0) glycoform. The gene for codon-optimized trastuzumab (TMab) was then introduced into PhytoRice® through Agrobacterium co-cultivation. Selected cell lines were suspension cultured, and TMab secreted from cells was purified from the cultured media. The amino acid sequence of the TMab produced by PhytoRice® (P-TMab) was identical to that of TMab. The inhibitory effect of P-TMab on the proliferation of the BT-474 cancer cell line was significantly enhanced at concentrations above 1 μg/mL (****P < 0.0001). P-TMab bound to a FcγRIIIa variant, FcγRIIIa-F158, more than 2.7 times more effectively than TMab. The ADCC efficacy of P-TMab against Jurkat cells was 2.6 times higher than that of TMab in an in vitro ADCC assay. Furthermore, P-TMab demonstrated efficient tumour uptake with less liver uptake compared to TMab in a xenograft assay using the BT-474 mouse model. These results suggest that the glycoengineered PhytoRice® could be an alternative platform for mAb production compared to current CHO cells, and P-TMab has a novel and enhanced efficacy compared to TMab.
{"title":"Enhanced efficacy of glycoengineered rice cell-produced trastuzumab","authors":"Jun-Hye Shin, Sera Oh, Mi-Hwa Jang, Seok-Yong Lee, Chanhong Min, Young-Jae Eu, Hilal Begum, Jong-Chan Kim, Gap Ryol Lee, Han-Bin Oh, Matthew J. Paul, Julian K.-C. Ma, Ho-Shin Gwak, Hyewon Youn, Seong-Ryong Kim","doi":"10.1111/pbi.14429","DOIUrl":"10.1111/pbi.14429","url":null,"abstract":"<p>For several decades, a plant-based expression system has been proposed as an alternative platform for the production of biopharmaceuticals including therapeutic monoclonal antibodies (mAbs), but the immunogenicity concerns associated with plant-specific N-glycans attached in plant-based biopharmaceuticals has not been completely solved. To eliminate all plant-specific N-glycan structure, eight genes involved in plant-specific N-glycosylation were mutated in rice (<i>Oryza sativa</i>) using the CRISPR/Cas9 system. The glycoengineered cell lines, PhytoRice®, contained a predominant GnGn (G0) glycoform. The gene for codon-optimized trastuzumab (TMab) was then introduced into PhytoRice® through <i>Agrobacterium</i> co-cultivation. Selected cell lines were suspension cultured, and TMab secreted from cells was purified from the cultured media. The amino acid sequence of the TMab produced by PhytoRice® (P-TMab) was identical to that of TMab. The inhibitory effect of P-TMab on the proliferation of the BT-474 cancer cell line was significantly enhanced at concentrations above 1 μg/mL (****<i>P</i> < 0.0001). P-TMab bound to a FcγRIIIa variant, FcγRIIIa-F158, more than 2.7 times more effectively than TMab. The ADCC efficacy of P-TMab against Jurkat cells was 2.6 times higher than that of TMab in an <i>in vitro</i> ADCC assay. Furthermore, P-TMab demonstrated efficient tumour uptake with less liver uptake compared to TMab in a xenograft assay using the BT-474 mouse model. These results suggest that the glycoengineered PhytoRice® could be an alternative platform for mAb production compared to current CHO cells, and P-TMab has a novel and enhanced efficacy compared to TMab.</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3068-3081"},"PeriodicalIF":10.1,"publicationDate":"2024-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14429","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141625531","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
<p>The crude oil crisis causes an increasing demand of renewable energy, among which, bioethanol is considered the cleanest and renewable liquid fuel alternative to fossil fuel (An Tran <i>et al</i>., <span>2019</span>). Bioethanol was mostly produced from sugarcane and corn, which violates vigorously against the world's food security. Alternatively, efforts have been made to produce bioethanol from non-food lignocellulose biomass, for example poplar, switchgrass and crop stover (An Tran <i>et al</i>., <span>2019</span>; Cai <i>et al</i>., <span>2016</span>; Fu <i>et al</i>., <span>2011</span>). Among which, corn stover is the most prevalent carbon-neutral lignocellulosic feedstock for the production of bioethanol although it is far from well utilized in bioethanol industry (Torres <i>et al</i>., <span>2014</span>). Lignocellulose is mainly composed of lignin, cellulose and hemicellulose. As lignin can reduce the availability of cellulose, pretreatment of corn stover by chemical reagents like diluted acids (4% H<sub>2</sub>SO<sub>4</sub>) to degrade lignin is a critical step prior to cellulose hydrolysis and fermentation (Figure 1a). However, the residual acids and the released phenolics and furfural compounds during pretreatment could inhibit the growth of microorganism in fermentation process thus decrease the bioethanol production efficiency and increase the processing cost (Rosales-Calderon and Arantes, <span>2019</span>; Zhao <i>et al</i>., <span>2013</span>). Therefore, lignin becomes the main barrier of ethanol production from lignocellulose, and searching for the lignin-reduced maize genetic materials is critical for the utilization of lignocellulosic biomass of corn stover in the production of bioethanol (Figure 1a).</p><p>Here, we screened a series of maize mutants potentially defective in lignin biosynthesis. Since <i>NST1</i> and <i>NST2</i> are key transcriptional regulators of secondary cell wall biogenesis in Arabidopsis (Mitsuda <i>et al</i>., <span>2005</span>), we obtained the mutants of their maize homologue genes (Figure S1) and evaluated their potential utilization in bioethanol production. Among them, <i>ZmNST2</i> express in all tissues including leaf, internode, root and shoot, with the highest expression detected in immature leaves (Figure S2). There were two G-to-A mutations that produce the premature stop codon in the second exon of <i>ZmNST2</i> in <i>zmnst2-1</i> and <i>zmnst2-2</i> mutant, respectively (Figure 1b). Both mutants are not morphologically different with the wild-type (WT) B73 except that the mutant leaves are softer and the mutants are slightly (4.86–6.63%) shorter (Figure S3a). The stem thickness, stem strength and dry biomass weight of the two mutants are not significantly different from the WT B73 (Figure S3b–e). We performed allelic test by crossing the two mutants to generate <i>zmnst2-1</i>/<i>zmnst2-2</i> F<sub>1</sub> plants, and the same soft-leaf phenotype was observed for the single mutants an
{"title":"Knockout of ZmNST2 promotes bioethanol production from corn stover","authors":"Ying Wang, Ye Xing, Xinyu Yang, Yanwen Yu, Jiankun Li, Chenyang Zhao, Mengyu Yuan, Weili Huang, Yue Yin, Guohui Liu, Yuqing Sun, Haochuan Li, Jihua Tang, Qin Zhang, Mingyue Gou","doi":"10.1111/pbi.14432","DOIUrl":"10.1111/pbi.14432","url":null,"abstract":"<p>The crude oil crisis causes an increasing demand of renewable energy, among which, bioethanol is considered the cleanest and renewable liquid fuel alternative to fossil fuel (An Tran <i>et al</i>., <span>2019</span>). Bioethanol was mostly produced from sugarcane and corn, which violates vigorously against the world's food security. Alternatively, efforts have been made to produce bioethanol from non-food lignocellulose biomass, for example poplar, switchgrass and crop stover (An Tran <i>et al</i>., <span>2019</span>; Cai <i>et al</i>., <span>2016</span>; Fu <i>et al</i>., <span>2011</span>). Among which, corn stover is the most prevalent carbon-neutral lignocellulosic feedstock for the production of bioethanol although it is far from well utilized in bioethanol industry (Torres <i>et al</i>., <span>2014</span>). Lignocellulose is mainly composed of lignin, cellulose and hemicellulose. As lignin can reduce the availability of cellulose, pretreatment of corn stover by chemical reagents like diluted acids (4% H<sub>2</sub>SO<sub>4</sub>) to degrade lignin is a critical step prior to cellulose hydrolysis and fermentation (Figure 1a). However, the residual acids and the released phenolics and furfural compounds during pretreatment could inhibit the growth of microorganism in fermentation process thus decrease the bioethanol production efficiency and increase the processing cost (Rosales-Calderon and Arantes, <span>2019</span>; Zhao <i>et al</i>., <span>2013</span>). Therefore, lignin becomes the main barrier of ethanol production from lignocellulose, and searching for the lignin-reduced maize genetic materials is critical for the utilization of lignocellulosic biomass of corn stover in the production of bioethanol (Figure 1a).</p><p>Here, we screened a series of maize mutants potentially defective in lignin biosynthesis. Since <i>NST1</i> and <i>NST2</i> are key transcriptional regulators of secondary cell wall biogenesis in Arabidopsis (Mitsuda <i>et al</i>., <span>2005</span>), we obtained the mutants of their maize homologue genes (Figure S1) and evaluated their potential utilization in bioethanol production. Among them, <i>ZmNST2</i> express in all tissues including leaf, internode, root and shoot, with the highest expression detected in immature leaves (Figure S2). There were two G-to-A mutations that produce the premature stop codon in the second exon of <i>ZmNST2</i> in <i>zmnst2-1</i> and <i>zmnst2-2</i> mutant, respectively (Figure 1b). Both mutants are not morphologically different with the wild-type (WT) B73 except that the mutant leaves are softer and the mutants are slightly (4.86–6.63%) shorter (Figure S3a). The stem thickness, stem strength and dry biomass weight of the two mutants are not significantly different from the WT B73 (Figure S3b–e). We performed allelic test by crossing the two mutants to generate <i>zmnst2-1</i>/<i>zmnst2-2</i> F<sub>1</sub> plants, and the same soft-leaf phenotype was observed for the single mutants an","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3099-3101"},"PeriodicalIF":10.1,"publicationDate":"2024-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14432","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141615444","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Bingting Wang, Zhian Wang, Ye Tang, Naiqin Zhong, Jiahe Wu
The Arabidopsis BLADE-ON-PETIOLE (BOP) genes are primarily known for their roles in regulating leaf and floral patterning. However, the broader functions of BOPs in regulating plant traits remain largely unexplored. In this study, we investigated the role of the Gossypium hirsutum BOP1 gene in the regulation of fibre length and plant height through the brassinosteroid (BR) signalling pathway. Transgenic cotton plants overexpressing GhBOP1 display shorter fibre lengths and reduced plant height compared to the wild type. Conversely, GhBOP1 knockdown led to increased plant height and longer fibre, indicating a connection with phenotypes influenced by the BR pathway. Our genetic evidence supports the notion that GhBOP1 regulates fibre length and plant height in a GhBES1-dependent manner, with GhBES1 being a major transcription factor in the BR signalling pathway. Yeast two-hybrid, luciferase complementation assay and pull-down assay results demonstrated a direct interaction between GhBOP1 and GhSUMO1, potentially forming protein complexes with GhBES1. In vitro and in vivo SUMOylation analyses revealed that GhBOP1 functions in an E3 ligase-like manner to mediate GhBES1 SUMOylation and subsequent degradation. Therefore, our study not only uncovers a novel mechanism of GhBES1 SUMOylation but also provides significant insights into how GhBOP1 regulates fibre length and plant height by controlling GhBES1 accumulation.
{"title":"Cotton BOP1 mediates SUMOylation of GhBES1 to regulate fibre development and plant architecture","authors":"Bingting Wang, Zhian Wang, Ye Tang, Naiqin Zhong, Jiahe Wu","doi":"10.1111/pbi.14428","DOIUrl":"10.1111/pbi.14428","url":null,"abstract":"<p>The Arabidopsis <i>BLADE-ON-PETIOLE</i> (<i>BOP</i>) genes are primarily known for their roles in regulating leaf and floral patterning. However, the broader functions of <i>BOPs</i> in regulating plant traits remain largely unexplored. In this study, we investigated the role of the <i>Gossypium hirsutum BOP1</i> gene in the regulation of fibre length and plant height through the brassinosteroid (BR) signalling pathway. Transgenic cotton plants overexpressing <i>GhBOP1</i> display shorter fibre lengths and reduced plant height compared to the wild type. Conversely, GhBOP1 knockdown led to increased plant height and longer fibre, indicating a connection with phenotypes influenced by the BR pathway. Our genetic evidence supports the notion that GhBOP1 regulates fibre length and plant height in a GhBES1-dependent manner, with GhBES1 being a major transcription factor in the BR signalling pathway. Yeast two-hybrid, luciferase complementation assay and pull-down assay results demonstrated a direct interaction between GhBOP1 and GhSUMO1, potentially forming protein complexes with GhBES1. <i>In vitro</i> and <i>in vivo</i> SUMOylation analyses revealed that GhBOP1 functions in an E3 ligase-like manner to mediate GhBES1 SUMOylation and subsequent degradation. Therefore, our study not only uncovers a novel mechanism of GhBES1 SUMOylation but also provides significant insights into how GhBOP1 regulates fibre length and plant height by controlling GhBES1 accumulation.</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3054-3067"},"PeriodicalIF":10.1,"publicationDate":"2024-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14428","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141602897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
<p>Approximately one-third of the total annual food production in the world is lost owing to pests, diseases and weeds. Therefore, the challenges posed by crop losses and population growth have emphasized the need for better breeding techniques (FAO <i>et al</i>., <span>2023</span>). Practical experience has demonstrated that the utilization of existing resistance genes to breed and cultivate herbicide- and pest-resistant rice cultivars is the most economical, safe and effective method for preventing and controlling weeds and pests (Zhang, <span>2007</span>).</p><p>The incorporation of a single or few resistance genes during rice breeding is no longer adequate for in-demand production. In addition, hybridization and backcrossing involve a long breeding cycle, and the issue of linkage drag may occur. The multi-gene transformation strategy can be utilized for the rapid and accurate incorporation of multiple resistance genes into rice (Zhu <i>et al</i>., <span>2017</span>). The fact that a trade-off between growth and defence generally exists in crops is universally accepted. Therefore, the overexpression of multi-resistance genes in rice causes considerable changes to the agronomic traits of crops, especially yield. The crop yield is positively correlated with the flowering stage within a certain range. For example, editing <i>Ehd1</i> or overexpressing <i>Ghd7</i> to appropriately extend the basic vegetative growth period of rice may be possible, and ultimately promote rice yield and quality (Eshed and Lippman, <span>2019</span>; Zhou <i>et al</i>., <span>2023</span>). This strategy is more effective for rice varieties with shorter growth periods. For some rice varieties with longer growth periods, we can use editing other yield related genes (grain type or grain weight), such as GS3 and GS5 (Ren <i>et al</i>., <span>2023</span>).</p><p>The herbicide resistance gene <i>I. variabilis-EPSPS*</i>, brown planthopper resistance genes <i>Bph14*</i> and <i>OsLecRK1*</i>, borer resistance gene <i>Cry1C*</i>, bacterial blight resistance gene <i>Xa23*</i> and blast resistance gene <i>Pi9*</i> are resistance gene resources in rice that have been extensively validated for use in rice breeding (Appendix S1). In our work, a highly efficient transgene system was used to construct an assembly of six resistance genes (about 26 Kb) mentioned earlier (380-6G) and <i>Ehd1</i> CRISPR/Cas9 editing vector (Cas9-<i>Ehd1</i>) (Figure 1a; Appendix S2 and S3). We expect to extend the basic vegetative growth period of multi-resistance gene transgenic rice by editing <i>Ehd1</i> to improve the agronomic traits (especially yield) and obtain a new multi-resistance and high-yield rice germplasm resource, termed MR&HY rice. We transformed two vectors, 380-6G and Cas9-<i>Ehd1</i>, into ZH11 rice varieties using <i>Agrobacterium</i>-mediated dual-strain transformation and screened using <i>glyphosate</i> and <i>hygromycin</i> simultaneously. When T<sub>0</sub> transgenic plants
{"title":"Development of a multi-resistance and high-yield rice variety using multigene transformation and gene editing","authors":"Changyan Li, Zaihui Zhou, Xinzhu Xiong, Chuanxu Li, Chuanhong Li, Enlong Shen, Jianyu Wang, Wenjun Zha, Bian Wu, Hao Chen, Lei Zhou, Yongjun Lin, Aiqing You","doi":"10.1111/pbi.14434","DOIUrl":"10.1111/pbi.14434","url":null,"abstract":"<p>Approximately one-third of the total annual food production in the world is lost owing to pests, diseases and weeds. Therefore, the challenges posed by crop losses and population growth have emphasized the need for better breeding techniques (FAO <i>et al</i>., <span>2023</span>). Practical experience has demonstrated that the utilization of existing resistance genes to breed and cultivate herbicide- and pest-resistant rice cultivars is the most economical, safe and effective method for preventing and controlling weeds and pests (Zhang, <span>2007</span>).</p><p>The incorporation of a single or few resistance genes during rice breeding is no longer adequate for in-demand production. In addition, hybridization and backcrossing involve a long breeding cycle, and the issue of linkage drag may occur. The multi-gene transformation strategy can be utilized for the rapid and accurate incorporation of multiple resistance genes into rice (Zhu <i>et al</i>., <span>2017</span>). The fact that a trade-off between growth and defence generally exists in crops is universally accepted. Therefore, the overexpression of multi-resistance genes in rice causes considerable changes to the agronomic traits of crops, especially yield. The crop yield is positively correlated with the flowering stage within a certain range. For example, editing <i>Ehd1</i> or overexpressing <i>Ghd7</i> to appropriately extend the basic vegetative growth period of rice may be possible, and ultimately promote rice yield and quality (Eshed and Lippman, <span>2019</span>; Zhou <i>et al</i>., <span>2023</span>). This strategy is more effective for rice varieties with shorter growth periods. For some rice varieties with longer growth periods, we can use editing other yield related genes (grain type or grain weight), such as GS3 and GS5 (Ren <i>et al</i>., <span>2023</span>).</p><p>The herbicide resistance gene <i>I. variabilis-EPSPS*</i>, brown planthopper resistance genes <i>Bph14*</i> and <i>OsLecRK1*</i>, borer resistance gene <i>Cry1C*</i>, bacterial blight resistance gene <i>Xa23*</i> and blast resistance gene <i>Pi9*</i> are resistance gene resources in rice that have been extensively validated for use in rice breeding (Appendix S1). In our work, a highly efficient transgene system was used to construct an assembly of six resistance genes (about 26 Kb) mentioned earlier (380-6G) and <i>Ehd1</i> CRISPR/Cas9 editing vector (Cas9-<i>Ehd1</i>) (Figure 1a; Appendix S2 and S3). We expect to extend the basic vegetative growth period of multi-resistance gene transgenic rice by editing <i>Ehd1</i> to improve the agronomic traits (especially yield) and obtain a new multi-resistance and high-yield rice germplasm resource, termed MR&HY rice. We transformed two vectors, 380-6G and Cas9-<i>Ehd1</i>, into ZH11 rice varieties using <i>Agrobacterium</i>-mediated dual-strain transformation and screened using <i>glyphosate</i> and <i>hygromycin</i> simultaneously. When T<sub>0</sub> transgenic plants","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3118-3120"},"PeriodicalIF":10.1,"publicationDate":"2024-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14434","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141602898","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
<p>Rice is a primary food crop, and its yield is threatened by biotic and abiotic stresses. <i>Xanthomonas oryzae</i> pv. <i>oryzae</i> (<i>Xoo</i>) causes bacterial blight, a chief bacterial disease of rice. <i>Xoo</i> infects rice depending on its transcriptional activation-like effectors (TALEs), which specifically target effector binding elements (EBEs) in the promoter of host susceptibility (<i>S</i>) genes and regulate <i>S</i> genes' expression for disease development. Editing <i>S</i> gene EBEs is an efficient approach for engineering disease-resistant rice (Oliva <i>et al</i>., <span>2019</span>; Xu <i>et al</i>., <span>2019</span>). Cold stress is a major abiotic factor that limits rice growth and productivity (Liu <i>et al</i>., <span>2019</span>). Therefore, engineering rice resilience to biotic and abiotic stresses is a powerful strategy to enhance rice yield. Here, we precisely edited an <i>S</i> gene EBE to engineer an elite rice variety exhibiting broad-spectrum resistance to <i>Xoo</i> and to enhanced cold tolerance.</p><p><i>OsTFX1</i> is an <i>S</i> gene targeted by the major <i>Xoo</i> TALE (Römer <i>et al</i>., <span>2010</span>; Sugio <i>et al</i>., <span>2007</span>). Analysing TALEs of <i>Xoo</i> whose genomic sequences are available, it was found that all <i>Xoo</i> strains contain a TALE that targets <i>OsTFX1</i> EBE (Figure S1) and activates <i>OsTFX1</i> expression (Yuan <i>et al</i>., <span>2016</span>), suggesting that <i>OsTFX1</i> is a <i>Xoo</i>-dependent major <i>S</i> gene. Comparing <i>OsTFX1</i> sequence in 3339 rice accessions from the RiceVarMap database, its EBE was found to have a unique sequence (Figure S2), indicating that there were no natural resistant alleles of <i>OsTFX1</i> for breeding. Therefore, we designed sgRNA specifically targeting <i>OsTFX1</i> EBE and generated the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants via CRISPR/Cas9-mediated mutagenesis. By screening 34 hygromycin-resistant independent lines by Sanger sequencing, we identified five types of <i>OsTFX1</i><sup><i>ebe</i></sup> mutants (Figure 1A). These <i>OsTFX1</i><sup><i>ebe</i></sup> mutants harbouring none off-target events were backcrossed with wild type (WT) and transgene-free plants were generated for analysis (Figure S3). The <i>OsTFX1</i><sup><i>ebe</i></sup> mutants exhibited enhanced resistance to a set of <i>Xoo</i> than WT (Figure 1B,C; Figure S4). <i>OsTFX1</i> did not respond to <i>Xoo</i> infection in the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants (Figure 1D), suggesting that EBE-edited <i>OsTFX1</i> had attenuated induction to <i>Xoo</i>, causing the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants to exhibit broad-spectrum resistance.</p><p>Interestingly, <i>OsTFX1</i> has significantly higher expression in <i>OsTFX1</i><sup><i>ebe</i></sup> mutants than in WT (Figure 1E). Diversification of transcription factor-targeted regulatory elements and modification of DNA methylation at the promoter can alter gene expression (Zhu
{"title":"Precision editing of a susceptibility gene promoter to alter its methylation modification for engineering rice resilience to biotic and abiotic stresses","authors":"Jingjing Tian, Hang Zhang, Shuxin Li, Yongjun Lin, Lizhong Xiong, Meng Yuan","doi":"10.1111/pbi.14430","DOIUrl":"10.1111/pbi.14430","url":null,"abstract":"<p>Rice is a primary food crop, and its yield is threatened by biotic and abiotic stresses. <i>Xanthomonas oryzae</i> pv. <i>oryzae</i> (<i>Xoo</i>) causes bacterial blight, a chief bacterial disease of rice. <i>Xoo</i> infects rice depending on its transcriptional activation-like effectors (TALEs), which specifically target effector binding elements (EBEs) in the promoter of host susceptibility (<i>S</i>) genes and regulate <i>S</i> genes' expression for disease development. Editing <i>S</i> gene EBEs is an efficient approach for engineering disease-resistant rice (Oliva <i>et al</i>., <span>2019</span>; Xu <i>et al</i>., <span>2019</span>). Cold stress is a major abiotic factor that limits rice growth and productivity (Liu <i>et al</i>., <span>2019</span>). Therefore, engineering rice resilience to biotic and abiotic stresses is a powerful strategy to enhance rice yield. Here, we precisely edited an <i>S</i> gene EBE to engineer an elite rice variety exhibiting broad-spectrum resistance to <i>Xoo</i> and to enhanced cold tolerance.</p><p><i>OsTFX1</i> is an <i>S</i> gene targeted by the major <i>Xoo</i> TALE (Römer <i>et al</i>., <span>2010</span>; Sugio <i>et al</i>., <span>2007</span>). Analysing TALEs of <i>Xoo</i> whose genomic sequences are available, it was found that all <i>Xoo</i> strains contain a TALE that targets <i>OsTFX1</i> EBE (Figure S1) and activates <i>OsTFX1</i> expression (Yuan <i>et al</i>., <span>2016</span>), suggesting that <i>OsTFX1</i> is a <i>Xoo</i>-dependent major <i>S</i> gene. Comparing <i>OsTFX1</i> sequence in 3339 rice accessions from the RiceVarMap database, its EBE was found to have a unique sequence (Figure S2), indicating that there were no natural resistant alleles of <i>OsTFX1</i> for breeding. Therefore, we designed sgRNA specifically targeting <i>OsTFX1</i> EBE and generated the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants via CRISPR/Cas9-mediated mutagenesis. By screening 34 hygromycin-resistant independent lines by Sanger sequencing, we identified five types of <i>OsTFX1</i><sup><i>ebe</i></sup> mutants (Figure 1A). These <i>OsTFX1</i><sup><i>ebe</i></sup> mutants harbouring none off-target events were backcrossed with wild type (WT) and transgene-free plants were generated for analysis (Figure S3). The <i>OsTFX1</i><sup><i>ebe</i></sup> mutants exhibited enhanced resistance to a set of <i>Xoo</i> than WT (Figure 1B,C; Figure S4). <i>OsTFX1</i> did not respond to <i>Xoo</i> infection in the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants (Figure 1D), suggesting that EBE-edited <i>OsTFX1</i> had attenuated induction to <i>Xoo</i>, causing the <i>OsTFX1</i><sup><i>ebe</i></sup> mutants to exhibit broad-spectrum resistance.</p><p>Interestingly, <i>OsTFX1</i> has significantly higher expression in <i>OsTFX1</i><sup><i>ebe</i></sup> mutants than in WT (Figure 1E). Diversification of transcription factor-targeted regulatory elements and modification of DNA methylation at the promoter can alter gene expression (Zhu","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3082-3084"},"PeriodicalIF":10.1,"publicationDate":"2024-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14430","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141602899","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Arturo Ortega, Kyungyong Seong, Alex Schultink, Daniela Paula de Toledo Thomazella, Eunyoung Seo, Elaine Zhang, Julie Pham, Myeong-Je Cho, Douglas Dahlbeck, Jacqueline Warren, Gerald V. Minsavage, Jeffrey B. Jones, Edgar Sierra-Orozco, Samuel F. Hutton, Brian Staskawicz
<p>Bacterial spot, caused by <i>Xanthomonas</i> species, is a devastating disease of tomato (<i>Solanum lycopersicum</i>) and pepper (<i>Capsicum annuum</i>) (Schwartz <i>et al</i>., <span>2015</span>). The recessively inherited resistance, <i>bacterial spot 5</i> (<i>bs5</i>), in pepper (hereafter referred to as <i>Cabs5</i>) can confer resistance against different <i>Xanthomonas</i> strains (Jones <i>et al</i>., <span>2002</span>). The <i>Cabs5</i> resistance is characterized by the absence of disease symptoms, faint chlorosis at the site of infection, and reduced bacterial growth. Remarkably, commercial pepper varieties containing the <i>bs5</i> allele show durable resistance, effectively impeding hypervirulent strain emergence in agricultural fields (Vallejos <i>et al</i>., <span>2010</span>).</p><p>The <i>CaBs5</i> gene, together with its paralog <i>CaBs5-like</i> (<i>CaBs5L</i>), has recently been cloned (Sharma <i>et al</i>., <span>2023</span>; Szabó <i>et al</i>., <span>2023</span>). <i>CaBs5</i> encodes a 92 amino acid long protein possessing a cysteine-rich transmembrane (CYSTM) domain, which is implicated in various biotic and abiotic responses. Typically, the CYSTM domain contains conserved residues composed of four consecutive cysteines, followed by two hydrophobic amino acids. A recent study suggested that Cabs5 mediating the resistance against bacterial spot lacks these two conserved leucine residues within the CYSTM domain (Szabó <i>et al</i>., <span>2023</span>).</p><p>Tomatoes and peppers are close relatives in the Solanaceae family and commonly susceptible to <i>Xanthomonas</i> infection. Based on the current findings in pepper, we hypothesized that modifying the ortholog of <i>CaBs5</i> in tomato could confer resistance against <i>Xanthomonas</i>. Consequently, putative <i>Bs5</i> (<i>SlBs5</i>) and <i>Bs5L</i> (<i>SlBs5L</i>) were identified in tomato based on homology to <i>CaBs5</i>. Both <i>SlBs5</i> and <i>SlBs5L</i> were located on chromosome 9 with the same head-to-head orientation as their pepper homologues on chromosome 3 (Figure 1a). Despite short and highly similar amino acid sequences of SlBs5 and SlBs5L (Figure 1b), the conserved synteny and gene order in pepper and tomato genomes allowed the assignment of orthology for <i>Bs5</i> and <i>Bs5L</i>.</p><p>The mechanism by which the double leucine deletion in <i>Cabs5</i> leads to resistance against <i>Xanthomonas</i> remains elusive (Figure 1b). Yet, this deletion in the conserved CYSTM domain could potentially impair CaBs5's native functionality (Abell and Mullen, <span>2011</span>). Following this assumption, we postulated that knocking out <i>SlBs5</i> would produce similar outcomes to <i>Cabs5</i>. We aimed to disrupt both SlBs5 and SlBs5L to prevent possible functional complementation by SlBs5L, given their greater amino acid sequence similarity compared to CaBs5 and CaBs5L (Figure 1b).</p><p>We constructed a binary vector for Cas9 and a single-guide RNA (sgRNA
{"title":"CRISPR/Cas9-mediated editing of Bs5 and Bs5L in tomato leads to resistance against Xanthomonas","authors":"Arturo Ortega, Kyungyong Seong, Alex Schultink, Daniela Paula de Toledo Thomazella, Eunyoung Seo, Elaine Zhang, Julie Pham, Myeong-Je Cho, Douglas Dahlbeck, Jacqueline Warren, Gerald V. Minsavage, Jeffrey B. Jones, Edgar Sierra-Orozco, Samuel F. Hutton, Brian Staskawicz","doi":"10.1111/pbi.14404","DOIUrl":"10.1111/pbi.14404","url":null,"abstract":"<p>Bacterial spot, caused by <i>Xanthomonas</i> species, is a devastating disease of tomato (<i>Solanum lycopersicum</i>) and pepper (<i>Capsicum annuum</i>) (Schwartz <i>et al</i>., <span>2015</span>). The recessively inherited resistance, <i>bacterial spot 5</i> (<i>bs5</i>), in pepper (hereafter referred to as <i>Cabs5</i>) can confer resistance against different <i>Xanthomonas</i> strains (Jones <i>et al</i>., <span>2002</span>). The <i>Cabs5</i> resistance is characterized by the absence of disease symptoms, faint chlorosis at the site of infection, and reduced bacterial growth. Remarkably, commercial pepper varieties containing the <i>bs5</i> allele show durable resistance, effectively impeding hypervirulent strain emergence in agricultural fields (Vallejos <i>et al</i>., <span>2010</span>).</p><p>The <i>CaBs5</i> gene, together with its paralog <i>CaBs5-like</i> (<i>CaBs5L</i>), has recently been cloned (Sharma <i>et al</i>., <span>2023</span>; Szabó <i>et al</i>., <span>2023</span>). <i>CaBs5</i> encodes a 92 amino acid long protein possessing a cysteine-rich transmembrane (CYSTM) domain, which is implicated in various biotic and abiotic responses. Typically, the CYSTM domain contains conserved residues composed of four consecutive cysteines, followed by two hydrophobic amino acids. A recent study suggested that Cabs5 mediating the resistance against bacterial spot lacks these two conserved leucine residues within the CYSTM domain (Szabó <i>et al</i>., <span>2023</span>).</p><p>Tomatoes and peppers are close relatives in the Solanaceae family and commonly susceptible to <i>Xanthomonas</i> infection. Based on the current findings in pepper, we hypothesized that modifying the ortholog of <i>CaBs5</i> in tomato could confer resistance against <i>Xanthomonas</i>. Consequently, putative <i>Bs5</i> (<i>SlBs5</i>) and <i>Bs5L</i> (<i>SlBs5L</i>) were identified in tomato based on homology to <i>CaBs5</i>. Both <i>SlBs5</i> and <i>SlBs5L</i> were located on chromosome 9 with the same head-to-head orientation as their pepper homologues on chromosome 3 (Figure 1a). Despite short and highly similar amino acid sequences of SlBs5 and SlBs5L (Figure 1b), the conserved synteny and gene order in pepper and tomato genomes allowed the assignment of orthology for <i>Bs5</i> and <i>Bs5L</i>.</p><p>The mechanism by which the double leucine deletion in <i>Cabs5</i> leads to resistance against <i>Xanthomonas</i> remains elusive (Figure 1b). Yet, this deletion in the conserved CYSTM domain could potentially impair CaBs5's native functionality (Abell and Mullen, <span>2011</span>). Following this assumption, we postulated that knocking out <i>SlBs5</i> would produce similar outcomes to <i>Cabs5</i>. We aimed to disrupt both SlBs5 and SlBs5L to prevent possible functional complementation by SlBs5L, given their greater amino acid sequence similarity compared to CaBs5 and CaBs5L (Figure 1b).</p><p>We constructed a binary vector for Cas9 and a single-guide RNA (sgRNA","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 10","pages":"2785-2787"},"PeriodicalIF":10.1,"publicationDate":"2024-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14404","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141597981","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Christoph Dockter, Søren Knudsen, Magnus Wohlfahrt Rasmussen, Birgitte Skadhauge, Birger Lindberg Møller
<p>In nature, genetic variation occurs in every population and results in the evolution of a diversity of new properties, some of which promote the survival of the species. To accelerate nature's evolution based on genetic diversity, plant breeders may induce additional mutations to raise the number of genetic variations increasing the chances to obtain varieties with new desired traits like improved nutritive quality, yields and resilience to biotic and abiotic stress factors. Induced mutagenesis based on chemical mutagens is considered non-GM and has been used in barley (<i>Hordeum vulgare</i>) for decades (Hansson <i>et al</i>., <span>2024</span>). Reverse genetic techniques including TILLING (Targeting Induced Local Lesions in Genomes) screening methodology and more recently TILLING-by-sequencing spinoffs are tools used to identify individual plant variants with the desired valuable genomic alterations. However, these tools are hampered by low mutation capacity.</p><p>TILLING is a PCR-based technique designed to detect mismatched single nucleotides in a target gene. In 2023, Szarejko and her research group in Poland published a thorough overview of the TILLING success stories within the last 20 years (Szurman-Zubrzycka <i>et al</i>., <span>2023</span>) including a description of the TILLING populations in different barley cultivars and landraces obtained following chemical mutagenesis (Figure 1a). The TILLING population sizes range between 1372 and 9600 individual plant variants. The mutation frequencies are individually chosen and dose-dependent (1/154–1/2500 Kbp). When multiplied (# of individuals × mutations per individual), the total number of mutations present in barley TILLING populations ranges between 10 and 100 million (Figure 1a). This may sound like a lot, but with a barley genome size of around 4300 Mbp (here, RGT Planet; Jayakodi <i>et al</i>., <span>2020</span>), less than 2% of the nucleotides in the entire population are mutated. This severely reduces the possibility to find a desired mutation in TILLING populations. The FIND-IT technology is a new approach overriding these constraints.</p><p>The FIND-IT technology was published in Science Advances in 2022 (Figure 1b) (Knudsen <i>et al</i>., <span>2022</span>) and provides an agile and high-throughput approach to screen unprecedented large size chemically induced variant populations. FIND-IT combines systematic sample pooling and splitting with high-sensitivity, droplet digital PCR (ddPCR)–based genotyping for targeted identification of desired traits at single-nucleotide resolution. The ddPCR technology is 1000-fold more sensitive than conventional PCR. The FIND-IT approach is applicable to any living organism that can be grown in the field or in culture. The experimental approach is outlined in detail in Knudsen <i>et al</i>., <span>2022</span> and illustrated schematically in Figure 1b. In total, more than 500 000 FIND-IT barley variant plants are today available for screen
{"title":"Just FIND-IT: Harnessing the true power of induced mutagenesis","authors":"Christoph Dockter, Søren Knudsen, Magnus Wohlfahrt Rasmussen, Birgitte Skadhauge, Birger Lindberg Møller","doi":"10.1111/pbi.14427","DOIUrl":"10.1111/pbi.14427","url":null,"abstract":"<p>In nature, genetic variation occurs in every population and results in the evolution of a diversity of new properties, some of which promote the survival of the species. To accelerate nature's evolution based on genetic diversity, plant breeders may induce additional mutations to raise the number of genetic variations increasing the chances to obtain varieties with new desired traits like improved nutritive quality, yields and resilience to biotic and abiotic stress factors. Induced mutagenesis based on chemical mutagens is considered non-GM and has been used in barley (<i>Hordeum vulgare</i>) for decades (Hansson <i>et al</i>., <span>2024</span>). Reverse genetic techniques including TILLING (Targeting Induced Local Lesions in Genomes) screening methodology and more recently TILLING-by-sequencing spinoffs are tools used to identify individual plant variants with the desired valuable genomic alterations. However, these tools are hampered by low mutation capacity.</p><p>TILLING is a PCR-based technique designed to detect mismatched single nucleotides in a target gene. In 2023, Szarejko and her research group in Poland published a thorough overview of the TILLING success stories within the last 20 years (Szurman-Zubrzycka <i>et al</i>., <span>2023</span>) including a description of the TILLING populations in different barley cultivars and landraces obtained following chemical mutagenesis (Figure 1a). The TILLING population sizes range between 1372 and 9600 individual plant variants. The mutation frequencies are individually chosen and dose-dependent (1/154–1/2500 Kbp). When multiplied (# of individuals × mutations per individual), the total number of mutations present in barley TILLING populations ranges between 10 and 100 million (Figure 1a). This may sound like a lot, but with a barley genome size of around 4300 Mbp (here, RGT Planet; Jayakodi <i>et al</i>., <span>2020</span>), less than 2% of the nucleotides in the entire population are mutated. This severely reduces the possibility to find a desired mutation in TILLING populations. The FIND-IT technology is a new approach overriding these constraints.</p><p>The FIND-IT technology was published in Science Advances in 2022 (Figure 1b) (Knudsen <i>et al</i>., <span>2022</span>) and provides an agile and high-throughput approach to screen unprecedented large size chemically induced variant populations. FIND-IT combines systematic sample pooling and splitting with high-sensitivity, droplet digital PCR (ddPCR)–based genotyping for targeted identification of desired traits at single-nucleotide resolution. The ddPCR technology is 1000-fold more sensitive than conventional PCR. The FIND-IT approach is applicable to any living organism that can be grown in the field or in culture. The experimental approach is outlined in detail in Knudsen <i>et al</i>., <span>2022</span> and illustrated schematically in Figure 1b. In total, more than 500 000 FIND-IT barley variant plants are today available for screen","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3051-3053"},"PeriodicalIF":10.1,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14427","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141562224","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Decades of studies have shown that Bt corn, by reducing insect damage, has lower levels of mycotoxins (fungal toxins), such as aflatoxin and fumonisin, than conventional corn. We used crop insurance data to infer that this benefit from Bt crops extends to reducing aflatoxin risk in peanuts: a non-Bt crop. In consequence, we suggest that any benefit–cost assessment of how transgenic Bt crops affect food safety should not be limited to assessing those crops alone; because the insect pest control offered by Bt crops affects the food safety profile of other crops grown nearby. Specifically, we found that higher Bt corn and Bt cotton planting rates in peanut-growing areas of the United States were associated with lower aflatoxin risk in peanuts as measured by aflatoxin-related insurance claims filed by peanut growers. Drought-related insurance claims were also lower: possibly due to Bt crops' suppression of insects that would otherwise feed on roots, rendering peanut plants more vulnerable to drought. These findings have implications for countries worldwide where policies allow Bt cotton but not Bt food crops to be grown: simply planting a Bt crop may reduce aflatoxin and drought stress in nearby food crops, resulting in a safer food supply through an inter-crop “halo effect.”
{"title":"Bt corn and cotton planting may benefit peanut growers by reducing aflatoxin risk","authors":"Jina Yu, David A. Hennessy, Felicia Wu","doi":"10.1111/pbi.14425","DOIUrl":"10.1111/pbi.14425","url":null,"abstract":"<p>Decades of studies have shown that Bt corn, by reducing insect damage, has lower levels of mycotoxins (fungal toxins), such as aflatoxin and fumonisin, than conventional corn. We used crop insurance data to infer that this benefit from Bt crops extends to reducing aflatoxin risk in peanuts: a non-Bt crop. In consequence, we suggest that any benefit–cost assessment of how transgenic Bt crops affect food safety should not be limited to assessing those crops alone; because the insect pest control offered by Bt crops affects the food safety profile of other crops grown nearby. Specifically, we found that higher Bt corn and Bt cotton planting rates in peanut-growing areas of the United States were associated with lower aflatoxin risk in peanuts as measured by aflatoxin-related insurance claims filed by peanut growers. Drought-related insurance claims were also lower: possibly due to Bt crops' suppression of insects that would otherwise feed on roots, rendering peanut plants more vulnerable to drought. These findings have implications for countries worldwide where policies allow Bt cotton but not Bt food crops to be grown: simply planting a Bt crop may reduce aflatoxin and drought stress in nearby food crops, resulting in a safer food supply through an inter-crop “halo effect.”</p>","PeriodicalId":221,"journal":{"name":"Plant Biotechnology Journal","volume":"22 11","pages":"3028-3036"},"PeriodicalIF":10.1,"publicationDate":"2024-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/pbi.14425","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141553830","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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