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| 1 | CRISPR/Cas9-mediated mutagenesis of VvMLO3 results in enhanced resistance to powdery mildew in grapevine(Vitis vinifera)显示文摘Grapevine(Vitis vinifera),one of the most economically important fruit crops in the world,suffers significant yield losses from powdery mildew,a major fungal disease caused by Erysiphe necator.In addition to suppressing host immunity,phytopathogens modulate host proteins termed susceptibility(S)factors to promote their proliferation in plants.In this study,CRISPR/Cas9(clustered regularly interspaced short palindromic repeats/CRISPR-associated 9)technology was used to enable the targeted mutagenesis of MLO(mildew resistance Locus O)family genes that are thought to serve as S factors for powdery mildew fungi.Small deletions or insertions were induced in one or both alleles of two grapevine MLO genes,VvMLO3 and VvMLO4,in the transgenic plantlets of the powdery mildew-susceptible cultivar Thompson Seedless.The editing efficiency achieved with different CRISPR/Cas9 constructs varied from 0 to 38.5%.Among the 20 VvMLO3/4-edited lines obtained,one was homozygous for a single mutation,three harbored biallelic mutations,seven were heterozygous for the mutations,and nine were chimeric,as indicated by the presence of more than two mutated alleles in each line.Six of the 20 VvMLO3/4-edited grapevine lines showed normal growth,while the remaining lines exhibited senescence-like chlorosis and necrosis.Importantly,four VvMLO3-edited lines showed enhanced resistance to powdery mildew,which was associated with host cell death,cell wall apposition(CWA)and H2O2 accumulation.Taken together,our results demonstrate that CRISPR/Cas9 genome-editing technology can be successfully used to induce targeted mutations in genes of interest to improve traits of economic importance,such as disease resistance in grapevines. | Dong-Yan Wan Ye Guo Yuan Cheng Yang Hu Shunyuan Xiao Yuejin Wang Ying-Qiang Wen | 2020 | Horticulture Research2020，7，1： | 4 |
| 2 | CRISPR/Cas技术在木本植物改良中的应用显示文摘木本植物育种周期长、种质资源匮乏,已成为限制木本植物种质创新的主要因素。CRISPR/Cas系统是近年来发展起来的能够实现对基因组进行精准定点编辑的技术,具有操作简单、周期短、效率高等优点,可实现基因的敲除、插入、替换等目的,从而精确地引入目标性状。目前,该技术已在多种家畜、昆虫、作物中得到了成功应用,对大量性状进行了改良,实现了种质资源的原始创新,大大缩短了育种年限。CRISPR/Cas技术的产生为木本植物的遗传改良提供了契机。笔者对CRISPR/Cas技术在杨树(Populus spp.)、苹果(Malus spp.)、柑橘(Citrus spp.)、葡萄(Vitis vinifera)、木薯(Cassava spp.)等木本植物育种中的应用进行了总结,该技术实现了T0代木本植物优良基因型的固定,在生长、材性、抗病性、抗旱性等性状上得到了明显改善,加快了木本植物育种进程,提高了育种效率。但该技术在木本植物中的应用尚处于起步阶段,还存在脱靶率高、编辑效率低、纯合突变少等问题。基于此,对CRISPR/Cas技术在木本植物中的应用前景进行了展望,以期为木本植物基因功能研究及品种改良提供有益参考。 | 侯静 毛金燕 翟惠 王洁 尹佟明 | 2021 | 南京林业大学学报(自然科学版)2021，45，6： | 3 |
| 3 | Breeding next generation tree fruits: technical and legal challenges显示文摘The new plant breeding technologies(NPBTs)have recently emerged as powerful tools in the context of‘green’biotechnologies.They have wide potential compared to classical genetic engineering and they are attracting the interest of politicians,stakeholders and citizens due to the revolutionary impact they may have on agriculture.Cisgenesis and genome editing potentially allow to obtain pathogen-resistant plants or plants with enhanced qualitative traits by introducing or disrupting specific genes in shorter times compared to traditional breeding programs and by means of minimal modifications in the plant genome.Grapevine,the most important fruit crop in the world from an economical point of view,is a peculiar case for NPBTs because of the load of cultural aspects,varietal traditions and consumer demands,which hinder the use of classical breeding techniques and,furthermore,the application of genetic engineering to wine grape cultivars.Here we explore the technical challenges which may hamper the application of cisgenesis and genome editing to this perennial plant,in particular focusing on the bottlenecks of the Agrobacterium-mediated gene transfer.In addition,strategies to eliminate undesired sequences from the genome and to choose proper target sites are discussed in light of peculiar features of this species.Furthermore is reported an update of the international legislative frameworks regulating NPBT products which shows conflicting positions and,in the case of the European Union,a prolonged lack of regulation. | Lorenza Dalla Costa Mickael Malnoy Ivana Gribaudo | 2017 | Horticulture Research2017，4，1： | 1 |
| 4 | Comparative proteomic analysis of cucumber powdery mildew resistance between a singlesegment substitution line and its recurrent parent显示文摘Powdery mildew(PM)is considered a major cause of yield losses and reduced quality in cucumber worldwide,but the molecular basis of PM resistance remains poorly understood.A segment substitution line,namely,SSL508-28,was developed with dominant PM resistance in the genetic background of PM-susceptible cucumber inbred line D8.The substituted segment contains 860 genes.An iTRAQ-based comparative proteomic technology was used to map the proteomes of PM-inoculated and untreated(control)D8 and SSL508-28.The number of differentially regulated proteins(DRPs)in SSL508-28 was almost three times higher than that in D8.Fourteen DRPs were located in the substituted segment interval.Comparative gene expression analysis revealed that nodulin-related protein 1(NRP1)may be a good candidate for PM resistance.Gene Ontology enrichment analysis showed that DRPs functioning in tetrapyrrole biosynthetic process,sulfur metabolic process and cell redox homeostasis were specifically enriched in the resistant line SSL508-28.DRPs categorized in the KEGG term photosynthesis increased in both lines upon PM infection,suggesting that the strategies used by cucumber may be different from those used by other crops to react to PM attacks at the initial stage.The measurement of hydrogen peroxide and superoxide anion production and net photosynthetic rate were consistent with the changes in protein abundance,suggesting that the proteomic results were reliable.There was a poor correlation between DRPs measured by iTRAQ and the corresponding gene expression changes measured by RNA-seq with the same experimental design.Taken together,these findings improve the understanding of the molecular mechanisms underlying the response of cucumber to PM infection. | Xuewen Xu Xueli Liu Yali Yan Wei Wang Kiros Gebretsadik Xiaohua Qi Qiang Xu Xuehao Chen | 2019 | Horticulture Research2019，6，1： | 1 |
| 5 | CmMLO17 and its partner CmKIC potentially support Alternaria alternata growth in Chrysanthemum morifolium显示文摘The Mildew Resistance Locus O(MLO)gene family has been investigated in many species.However,there are few studies on chrysanthemum MLO genes.We report in this study that CmMLO17 in Chrysanthemum morifolium was upregulated after Alternaria alternata infection.Silencing of CmMLO17 by artificial microRNA resulted in reduced susceptibility of chrysanthemum to A.alternata infection.Genes in the abscisic acid(ABA)and Ca 2+signaling pathways were upregulated in the CmMLO17-silenced line R20 compared to the wild-type plants.We speculated that CmMLO17-silenced plants had a faster and stronger defense response that was mediated by the ABA and Ca 2+signaling pathways,resulting in reduced susceptibility of chrysanthemum to A.alternata infection.In addition,a candidate gene,CmKIC,that may interact with CmMLO17 was discovered by the yeast two-hybrid assay.The interaction between CmMLO17 and CmKIC was confirmed using the yeast two-hybrid assay and bimolecular fluorescence complementation(BiFC)analysis.CmMLO17 and CmKIC were both located on the plasma membrane,and CmKIC was also located on the nucleus.CmKIC overexpression increased the susceptibility of chrysanthemum to A.alternata,whereas CmKIC silencing resulted in reduced susceptibility.Therefore,CmMLO17 and CmKIC may work together in C.morifolium to support the growth of A.alternata.The results of this study will provide insight into the potential function of MLO and improve the understanding of plant defense responses to necrotrophic pathogens. | Jingjing Xin Ye Liu Huiyun Li Sumei Chen Jiafu Jiang Aiping Song Weimin Fang Fadi Chen | 2021 | Horticulture Research2021，8，1： | 1 |
| 6 | Evolution of the MLO gene families in octoploid strawberry(Fragaria×ananassa)and progenitor diploid species identified potential genes for strawberry powdery mildew resistance显示文摘Powdery mildew(PM)caused by Podosphaera aphanis is a major fungal disease of cultivated strawberry.Mildew Resistance Locus O(MLO)is a gene family described for having conserved seven-transmembrane domains.Induced loss-of-function in specific MLO genes can confer durable and broad resistance against PM pathogens.However,the genomic structure and potential role of MLO genes for PM resistance have not been characterized yet in the octoploid cultivated strawberry.In the present study,MLO gene families were characterized in four diploid progenitor species(Fragaria vesca,F.iinumae,F.viridis,and F.nipponica)and octoploid cultivated(Fragaria×ananassa)strawberry,and potential sources of MLO-mediated susceptibility were identified.Twenty MLO sequences were identified in F.vesca and 68 identified in F.×ananassa.Phylogenetic analysis divided diploid and octoploid strawberry MLO genes into eight different clades,in which three FveMLO(MLO10,MLO17,and MLO20)and their twelve orthologs of FaMLO were grouped together with functionally characterized MLO genes conferring PM susceptibility.Copy number variations revealed differences in MLO composition among homoeologous chromosomes,supporting the distinct origin of each subgenome during the evolution of octoploid strawberry.Dissecting genomic sequence and structural variations in candidate FaMLO genes revealed their potential role associated with genetic controls and functionality in strawberry against PM pathogen.Furthermore,the gene expression profiling and RNAi silencing of putative FaMLO genes in response to the pathogen indicate the function in PM resistance.These results are a critical first step in understanding the function of strawberry MLO genes and will facilitate further genetic studies of PM resistance in cultivated strawberry. | Ronald R.Tapia Christopher R.Barbey Saket Chandra Kevin M.Folta Vance M.Whitaker Seonghee Lee | 2021 | Horticulture Research2021，8，1： | 1 |
| 7 | Can effectoromics and loss-of-susceptibility be exploited for improving Fusarium head blight resistance in wheat?显示文摘Bread wheat(Triticum aestiuum L.),which provides about 20%of daily calorie intake,is the most widely cultivated crop in the world,in terms of total area devoted to its cultivation.Therefore,even small increases in wheat yield can translate into large gains.Reducing the gap between actual and potential grain yield in wheat is a crucial task to feed the increasing world population.Fusarium head blight(FHB)caused by the pathogenic fungus Fusaium graminearum and related Fusarium species is one of the most devastating wheat diseases throughout the world.This disease reduces not only the yield but also the quality by contaminating the grain with mycotoxins harmful for humans,animals and the environment.In recent years,remarkable achievements attained in omics'technologies have not only provided new insights into understanding of processes involved in pathogenesis but also helped develop effective new tools for practical plant breeding.Sequencing of the genomes of various wheat patho gens,including F.graminearum,as well as those of bread and durum wheat and their wild relatives,together with advances made in transcriptomics and bioinformatics,has allowed the identification of candidate pathogen effectors and corresponding host resistance(R)and susceptibility(S)genes.However,so far,FHB effectors and wheat susceptibility genes/factors have been poorly studied.In this paper,we first briefly highlighted recent examples of improving resistance against pathogens via new techniques in different host species.We then propose effective strategies towards developing wheat cultivars with improved resistance to FHB.We hope that the article will spur discussions and interest among researchers about novel approaches with great potential for improving wheat against FHB. | Andrii Gorash Rita Armonienė Kemal Kazan | 2021 | The Crop Journal2021，9，1： | 0 |
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