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| 1 | A method for the production and expedient screening of CRISPR/Cas9-mediated non-transgenic mutant plants显示文摘Developing CRISPR/Cas9-mediated non-transgenic mutants in asexually propagated perennial crop plants is challenging but highly desirable.Here,we report a highly useful method using an Agrobacterium-mediated transient CRISPR/Cas9 gene expression system to create non-transgenic mutant plants without the need for sexual segregation.We have also developed a rapid,cost-effective,and high-throughput mutant screening protocol based on Illumina sequencing followed by high-resolution melting(HRM)analysis.Using tetraploid tobacco as a model species and the phytoene desaturase(PDS)gene as a target,we successfully created and expediently identified mutant plants,which were verified as tetra-allelic mutants.We produced pds mutant shoots at a rate of 47.5%from tobacco leaf explants,without the use of antibiotic selection.Among these pds plants,17.2%were confirmed to be non-transgenic,for an overall non-transgenic mutation rate of 8.2%.Our method is reliable and effective in creating non-transgenic mutant plants without the need to segregate out transgenes through sexual reproduction.This method should be applicable to many economically important,heterozygous,perennial crop species that are more difficult to regenerate. | Longzheng Chen Wei Li Lorenzo Katin-Grazzini Jing Ding Xianbin Gu Yanjun Li Tingting Gu Ren Wang Xinchun Lin Ziniu Deng Richard J.McAvoy Frederick G.Gmitter Jr. Zhanao Deng Yunde Zhao Yi Li | 2018 | Horticulture Research2018，5，1： | 12 |
| 2 | Reprogramming of a defense signaling pathway in rough lemon and sweet orange is a critical element of the early response to‘Candidatus Liberibacter asiaticus’显示文摘Huanglongbing(HLB)in citrus infected by Candidatus Liberibacter asiaticus(CLas)has caused tremendous losses to the citrus industry.No resistant genotypes have been identified in citrus species or close relatives.Among citrus varieties,rough lemon(Citrus jambhiri)has been considered tolerant due to its ability to produce a healthy flush of new growth after infection.The difference between tolerance and susceptibility is often defined by the speed and intensity of a plant’s response to a pathogen,especially early defense responses.RNA-seq data were collected from three biological replicates of CLas-and mock-inoculated rough lemon and sweet orange at week 0 and 7 following infection.Functional analysis of the differentially expressed genes(DEGs)indicated that genes involved in the mitogen activated protein kinase(MAPK)signaling pathway were highly upregulated in rough lemon.MAPK induces the transcription of WRKY and other transcription factors which potentially turn on multiple defense-related genes.A Subnetwork Enrichment Analysis further revealed different patterns of regulation of several functional categories,suggesting DEGs with different functions were subjected to reprogramming.In general,the amplitude of the expression of defense-related genes is much greater in rough lemon than in sweet orange.A quantitative disease resistance response may contribute to the durable tolerance level to HLB observed in rough lemon. | Qibin Yu Chunxian Chen Dongliang Du Ming Huang Jiqiang Yao Fahong Yu Ronald H Brlansky Frederick G.Gmitter Jr | 2017 | Horticulture Research2017，4，1： | 5 |
| 3 | 柑桔线虫抗性主基因座Tyr1的特异标记开发与遗传作图改进(英文)显示文摘本文利用先期从BAC文库获得的NBS-LRR类候选抗病基因克隆序列Pt8a和Pt9a,进一步开发与柑桔线虫抗性主效基因位点Tyr1连锁的分子标记。以Pt8a和Pt9a序列作探针,通过高密度克隆印迹杂交,从BAC文库筛选出200个以上的阳性克隆,以阳性克隆插入序列设计引物,对柑桔抗线虫材料和感线虫材料开展以PCR扩增为基础的集群分离分析,发现一部分克隆序列与柑桔线虫抗性主效基因位点Tyr1紧密连锁;再通过染色体步行测序,分别从3个克隆(7A4,4L17和29F20)获得3个完整的NBS-LRR类候选抗病基因序列。从此类序列开发更高特异性的分子标记,并在利用原有分子标记的基础上,对柑桔线虫抗性杂交后代群体(9145family)构建较高密度的遗传图谱;同时,将新开发的分子标记应用于柑桔衰退病抗性杂交后代群体(9401family),以初步估算柑桔线虫抗性主效基因位点Tyr1与柑桔衰退病抗性基因Ctv的遗传距离。本文利用先期从BAC文库获得的NBS-LRR类候选抗病基因克隆序列Pt8a和Pt9a,进一步开发与柑桔线虫抗性主效基因位点Tyr1连锁的分子标记。以Pt8a和Pt9a序列作探针,通过高密度克隆印迹杂交,从BAC文库筛选出200个以上的阳性克隆,以阳性克隆插入序列设计引物,对柑桔抗线虫材料和感线虫材料开展以PCR扩增为基础的集群分离分析,发现一部分克隆序列与柑桔线虫抗性主效基因位点Tyr1紧密连锁;再通过染色体步行测序,分别从3个克隆(7A4,4L17和29F20)获得3个完整的NBS-LRR类候选抗病基因序列。从此类序列开发更高特异性的分子标记,并在利用原有分子标记的基础上,对柑桔线虫抗性杂交后代群体(9145family)构建较高密度的遗传图谱;同时,将新开发的分子标记应用于柑桔衰退病抗性杂交后代群体(9401family),以初步估算柑桔线虫抗性主效基因位点Tyr1与柑桔衰退病抗性基因Ctv的遗传距离。 | 向旭 邓占鳌 郑启发 陈存贤 Frederick G.Gmitter Jr | 2009 | 分子植物育种2009，7，3： | 2 |
| 4 | Construction of citrus gene coexpression networks from microarray data using random matrix theory显示文摘After the sequencing of citrus genomes,gene function annotation is becoming a new challenge.Gene coexpression analysis can be employed for function annotation using publicly available microarray data sets.In this study,230 sweet orange(Citrus sinensis)microarrays were used to construct seven coexpression networks,including one condition-independent and six condition-dependent(Citrus canker,Huanglongbing,leaves,flavedo,albedo,and flesh)networks.In total,these networks contain 37633 edges among 6256 nodes(genes),which accounts for 52.11%measurable genes of the citrus microarray.Then,these networks were partitioned into functional modules using the Markov Cluster Algorithm.Significantly enriched Gene Ontology biological process terms and KEGG pathway terms were detected for 343 and 60 modules,respectively.Finally,independent verification of these networks was performed using another expression data of 371 genes.This study provides new targets for further functional analyses in citrus. | Dongliang Du Nidhi Rawat Zhanao Deng Fred G.Gmitter Jr. | 2015 | Horticulture Research2015，2，1： | 2 |
| 5 | Novel assembly strategy cracks open the mysteries of walnut genome evolution显示文摘High quality chromosome-scale assemblies from an interspecific hybrid between walnut and a wild relative reveal the persistence of asymmetric fractionation between the sub-genomes and suggest a late-Miocene origin for the genus Juglans. | G.Albert Wu Fred G.Gmitter | 2019 | Horticulture Research2019，6，1： | 1 |
| 6 | Changes in carbohydrate metabolism in Citrus sinensis infected with ‘Candidatus Liberibacter asiaticus’显示文摘 | J.Fan C.Chen R. H.Brlansky F. G.Gmitter Jr. Z.‐G.Li | 2010 | Plant Pathology2010，，6： | 1 |
| 7 | Mapping of QTLs and candidate genes associated with multiple phenotypic traits for Huanglongbing tolerance in citrus显示文摘Huanglongbing(HLB)is the most devastating disease for citrus worldwide.Candidatus Liberibacter asiaticus(C Las),vectored by Asian citrus psyllid(ACP,Diaphorina citri Kuwayama),is the most common pathogen causing the disease.Commercial citrus varieties are highly susceptible to HLB,whereas trifoliate orange(Poncirus trifoliata)is considered highly tolerant to HLB.An F1 segregating population and their parent trifoliate orange and sweet orange,which had been exposed to intense HLB pressure for three years,was evaluated for disease symptoms,ACP colonization,C Las titer and tree vigor repeatedly for two to three years.Trifoliate orange and sweet orange showed significant differences for most of the phenotypic traits,and the F1 population exhibited a large variation.A high-density SNP-based genetic map with 1402 markers was constructed for trifoliate orange,which exhibited high synteny and high coverage of its reference genome.A total of 26 quantitative trait locus(QTLs)were identified in four linkage groups LG-t6,LG-t7,LG-t8 and LG-t9,of which four QTL clusters exhibit a clear co-localization of QTLs associated with different traits.Through genome-wide analysis of gene expression in response to C Las infection in‘Flying Dragon’and‘Larger-Flower DPI-50-7’trifoliate orange,85 differentially expressed genes were found located within the QTL clusters.Among them,seven genes were classified as defense or immunity protein which exhibited the highest transcriptional change after C Las infection.Our results indicate a quantitative genetic nature of HLB tolerance and identified candidate genes that should be valuable for searching for genetic solutions to HLB through breeding or genetic engineering. | Ming Huang Mikeal L. Roose Qibin Yu Ed Stover David G. Hall Zhanao Deng Frederick G.Gmitter Jr. | 2023 | Horticultural Plant Journal2023，9，4： | 0 |
| 8 | Rationale for reconsidering current regulations restricting use of hybrids in orange juice显示文摘Huanglongbing(HLB)is a disease that has devastated the Florida citrus industry,threatens the entire U.S.citrus industry,and globally is rapidly spreading.Florida’s citrus production is 90%sweet orange,which is quite sensitive to HLB.The heavy reliance on sweet orange for Florida citrus production makes the industry especially vulnerable to diseases that are damaging to this type of citrus.Furthermore,90%of Florida oranges are used in producing orange juice that is defined by a federal regulation known as the“orange juice standard”,specifying that at least 90%of“orange juice”must be derived from Citrus sinensis.Genomic analyses definitively reveal that sweet orange is not a true species,but just one of many introgression hybrids of C.reticulata and C.maxima,with phenotypic diversity resulting from accumulated mutations in this single hybrid,the“sweet orange”.No other fruit industry is limited by law to such a narrow genetic base.Fortunately,there are new citrus hybrids displaying reduced sensitivity to HLB,and in some cases they produce juice,alone or in blends,that consumers would recognize as“orange juice”.Reconsidering current regulations on orange juice standards may permit use of such hybrids in“orange juice”,providing greater latitude for commercialization of these hybrids,leading to higher-quality orange juice and a more sustainable Florida orange juice industry. | Ed Stover Frederick G.Gmitter Jr Jude Grosser Elizabeth Baldwin Guohong Albert Wu Jinhe Bai Yu Wang Peter Chaires Juan Carlos Motamayor | 2020 | Horticulture Research2020，7，1： | 0 |
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