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WRKY11 up-regulated dirigent expression to enhance lignin/lignans accumulation in Lilium regale Wilson during response to Fusarium wilt

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Journal of Integrative Agriculture 2024年第8期 2703-2722页

作者： Jie Deng Zi'e Wang Wenyun Li Xiaohua Chen diqiu liu Faculty of Life Science and Technology Kunming University of Science and Technology

Lilium are highly economically valuable ornamental plants that are susceptible to Fusarium wilt caused by Fusarium oxysporum.Lilium regale Wilson, a wild lily native to China, is highly resistant to F. oxysporum.In this study, a WRKYtranscriptionfactor,WRKY11,wasisolatedfromL. regale,anditsfunctionduringtheinteractionbetween L.regaleandF.oxysporumwascharacterized.TheectopicexpressionofLrWRKY11intobaccoincreasedthe resistance to F. oxysporum, moreover, the transcriptome sequencing and UHPLC-MS/MS analysis indicated that the methyl salicylate and methyl jasmonate levels rose in LrWRKY11 transgenic tobacco, meanwhile, the expression of lignin/lignans biosynthesis-related genes including adirigent (DIR) was up-regulated.The lignin/lignans contents inLrWRKY11-transgenictobaccoalsosignificantlyincreasedcomparedwiththewild-typetobacco.Inaddition,theresistanceofL.regalescalesinwhichLrWRKY11expressionwassilencedbyRNAievidentlydecreased,andadditionally,theexpressionoflignin/lignansbiosynthesis-relatedgenesincludingLrDIR1wassignificantly suppressed.Therefore,LrDIR1anditspromoter(PLrDIR1)sequencecontainingtheW-boxelementwere isolated from L. regale.The interaction assay indicated that LrWRKY11 specifically bound to the W-box element in PLrDIR1andactivatedLrDIR1 expression.Additionally, β-glucuronidase activity in the transgenic tobacco coexpressingLrWRKY11/PLrDIR1-β-glucuronidase was higher than that in transgenic tobacco expressing PLrDIR1-β-glucuronidase alone.Furthermore, the ectopic expression of LrDIR1 in tobacco enhanced the resistance to F. oxysporum and increased the lignin/lignans accumulation.In brief, this study revealed that LrWRKY11 positively regulatedL.regaleresistancetoF.oxysporumthroughinteractionwithsalicylicacid/jasmonicacidsignaling pathways and modulating LrDIR1 expression to accumulate lignin/lignans.

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Prevalence and molecular epidemiology of equine piroplasmosis in China:a neglected tick-borne disease

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Science China(Life Sciences) 2022年第2期65卷 445-447页

作者： Kewei Chen Zhe Hu Jingkun Li Jingfei Wang diqiu liu Ting Qi Wei Guo Cheng Du Xiaojun Wang State Key Laboratory of Veterinary Biotechnology Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin 150069China Department of Biostatistics School of Public HealthHarbin Medical UniversityHarbin 150081China

Dear Editor,Equine piroplasmosis(EP)is an infectious,tick-borne dis-ease caused by the hemoprotozoan parasites Theileria equi(T.equi)and Babesia caballi(B.caballi).EP affects all wild and domestic equids,and clinical presentation is related to intravascular hemolysis and associated systemic illness(Scoles and Ueti,2015).This disease has considerable ve-terinary and economic impact on the equine industry worldwide,and is listed as a reportable disease by the World Organization for Animal Health(OIE).Importantly,a ne-gative EP test is mandatory for animals participating in equestrian sports and the horse trade.It is estimated that 90%of the global horse population resides in EP-endemic areas.Many risk factors are associated with the infection rate,in-cluding animal species,age,gender,breed,activities,and environmental factors(Onyiche et al.,2019).

关键词： horse epidemiology listed

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Equine ANP32 proteins support influenza A virus RNA polymerase activity

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Virologica Sinica 2023年第6期38卷 951-960页

作者： Yuan Zhang Xing Guo Mengmeng Yu liuke Sun Yuxing Qu Kui Guo Zhe Hu diqiu liu Haili Zhang Xiaojun Wang State Key Laboratory for Animal Disease Control and Prevention Harbin Veterinary Research InstituteThe Chinese Academy of Agricultural SciencesHarbin150069China State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases Key Laboratory for Zoonosis Research of the Ministry of EducationInstitute of Zoonosis and College of Veterinary MedicineJilin UniversityChangchun130062China

Host ANP32 family proteins are crucial for maintaining the activity of influenza RNA polymerase and play an important role in the cross-species transmission of influenza viruses.To date,the molecular properties of equine ANP32(eqANP32)protein are poorly understood,particularly the mechanisms that affect equine influenza virus(EIV)RNA polymerase activity.Here,we found that there are six alternative splicing variants of equine ANP32A(eqANP32A)with different levels of expression.Further studies showed that these six splicing variants of eqANP32A supported the activity of EIV RNA polymerase to varying degrees,with the variant eqANP32A_X2 having the highest expression abundance and exhibiting the highest support of polymerase activity.Sequence analysis demonstrated that the differences in the N-Cap regions of the six splicing variants significantly affected their N-terminal conformation,but did not affect their ability to bind RNA polymerase.We also demonstrated that there is only one transcript of eqANP32B,and that this transcript showed only very low support to the EIV RNA polymerase.This functional defect in eqANP32B is caused by the sequence of the 110–259 amino acids at its Cterminus.Our results indicated that it is the eqANP32A_X2 protein that mainly determines the efficiency of the EIV replication in horses.In conclusion,our study parsed the molecular properties of eqANP32 family proteins and revealed the sequence features of eqANP32A and eqANP32B,suggesting for the first time that the N-Cap region of ANP32A protein also plays an important role in supporting the activity of the influenza virus polymerase.

关键词： Equine influenza virus(EIV) Equine ANP32A Equine ANP32B RNA polymerase activity N-Cap domain

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Suitable internal control genes for qRT-PCR normalization in cotton fiber development and somatic embryogenesis

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Chinese Science Bulletin 2007年第22期52卷 3110-3117页

作者： TU LiLi ZHANG XianLong liu diqiu JIN ShuangXia CAO JingLin ZHU LongFu DENG FengLin TAN JiaFu ZHANG CunBin National Key Laboratory of Crop Genetic Improvement Huazhong Agricultural University Wuhan 430070 China

The mechanisms of cotton fiber development and somatic embryogenesis have been explored sys-tematically with microarray and suppression subtractive hybridization. Real-time RT-PCR provides the simultaneous measurement of gene expression in many different samples,with which the data from microarray or others can be confirmed in detail. To achieve accurate and reliable gene expression re-sults,normalization of real-time PCR data against one or several internal control genes is required,which should not fluctuate in different tissues during various stages of development. We assessed the gene expression of 7 frequently used housekeeping genes,including 18S rRNA,Histone3,UBQ7,Actin,Cyclophilin,Gbpolyubiquitin-1 and Gbpolyubiquitin-2,in a diverse set of 21 cotton samples. For fiber developmental series the expression of all housekeeping genes had the same down tendency after 17 DPA. But the expression of the AGP gene(arabinogalactan protein) that has high expression level at the later fiber development stage was up-regulated from 15 to 27 DPA. So the relative absolute quanti-fication should be an efficient and convenient method for the fiber developmental series. The expres-sion of nonfiber tissues series varied not so much against the fiber developmental series. And three best control genes Histone3,UBQ7 and Gbpolyubiquitin-1 have to be used in a combinated way to get better normalization.

关键词：棉花纤维胚胎测量模式

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PjFPS和Pjβ-AS双基因协同作用对珠子参皂苷生物合成的影响

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浙江农业学报 2022年第3期34卷 428-436页

作者：陈勤刘美佳刘迪秋曲媛崔秀明葛锋昆明理工大学生命科学与技术学院云南昆明650500 云南省三七资源可持续利用重点实验室云南昆明650500

法尼基焦磷酸合酶(PjFPS)和β-香树脂醇合酶(Pjβ-AS)是珠子参皂苷(Panax japonicus saponins,PJS)生物合成途径中可能的关键酶。通过在珠子参(P.japonicus)细胞中同时过表达PjFPS和Pjβ-AS基因,探讨PjFPS和Pjβ-AS对PJS生物合成的协同... 详细信息

法尼基焦磷酸合酶(PjFPS)和β-香树脂醇合酶(Pjβ-AS)是珠子参皂苷(Panax japonicus saponins,PJS)生物合成途径中可能的关键酶。通过在珠子参(P.japonicus)细胞中同时过表达PjFPS和Pjβ-AS基因,探讨PjFPS和Pjβ-AS对PJS生物合成的协同作用。结果表明,PjFPS和Pjβ-AS是PJS生物合成途径中的关键酶基因,对珠子参皂苷的生物合成具有调节作用。过表达PjFPS和Pjβ-AS的细胞系中,PJS合成途径相关的多个关键酶基因的表达水平有不同程度的提高,且PJS的含量最高为普通细胞系的2.4倍。虽然单独过表达PjFPS也可增加PJS的合成,但双基因(PjFPS和Pjβ-AS)协同调控PJS合成的效果更好。

关键词：三萜皂苷生物合成珠子参法尼基焦磷酸合酶 β-香树脂醇合酶

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交旅融合视域下旅游文创设计人才培养路径探索——以重庆交大毕业设计教学为例

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装饰 2020年第5期 130-131页

作者：刘第秋重庆交通大学艺术设计学院

本文基于交旅融合发展的时代需求,以学校交通类优势学科为基础支撑和先决条件,结合艺术设计学科特点,思索如何依托交通与旅游转型升级的契机,培养跨行业、具备综合素养的旅游文创设计人才。旨在通过产教融合和校企合作的方式,探讨如何... 详细信息

本文基于交旅融合发展的时代需求,以学校交通类优势学科为基础支撑和先决条件,结合艺术设计学科特点,思索如何依托交通与旅游转型升级的契机,培养跨行业、具备综合素养的旅游文创设计人才。旨在通过产教融合和校企合作的方式,探讨如何提升复合型艺术设计人才的市场针对性和核心竞争力。

关键词：交旅融合旅游文创文创设计人才培养

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苗族视觉文化与现代设计融合的探讨——贵州摆贝苗寨田野调查与实践

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装饰 2016年第3期 142-143页

作者：刘第秋重庆交通大学马蒂亚斯国际设计学院

本文针对苗族手工艺的现状、问题和趋势,借由对贵州边远苗族村寨的实地考察,进而思索如何将正在逐渐消失的苗族传统手工艺与现代设计融合,旨在探讨少数民族视觉文化与现代生活美学关联的途径。

关键词：手工艺苗族摆贝苗寨现代设计

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岷江百合多聚半乳糖醛酸酶抑制蛋白基因及其启动子的克隆与分析

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华北农学报 2022年第4期37卷 45-52页

作者：邓婕王瀚林李有玉王自娥陈晓华刘迪秋昆明理工大学生命科学与技术学院云南昆明650500

多聚半乳糖醛酸酶抑制蛋白(PGIPs)在植物抵御病原菌侵染的过程中具有重要的作用。从尖孢镰刀菌抗性百合野生种岷江百合中分离得到一个PGIP基因及其启动子。LrPGIP的开放阅读框长度为1008 bp,编码一个含有335个氨基酸残基的蛋白质,并具有... 详细信息

多聚半乳糖醛酸酶抑制蛋白(PGIPs)在植物抵御病原菌侵染的过程中具有重要的作用。从尖孢镰刀菌抗性百合野生种岷江百合中分离得到一个PGIP基因及其启动子。LrPGIP的开放阅读框长度为1008 bp,编码一个含有335个氨基酸残基的蛋白质,并具有7个富含亮氨酸重复结构域。LrPGIP与油棕、椰子、林烟草的PGIP同源性较高,分别为91%,94%,86%,且与单子叶植物海枣、粗柄象腿蕉、油棕、椰子的PGIP蛋白亲缘关系更近。LrPGIP在根、茎、叶、花、鳞片中均有表达,在根中的表达量最高,在鳞片中的表达量最低。LrPGIP的表达水平受尖孢镰刀菌的诱导,在接种尖孢镰刀菌后72 h表达量最高。植物信号分子水杨酸、茉莉酸甲酯、乙烯利、过氧化氢均诱导LrPGIP表达。LrPGIP受乙烯利诱导后的表达量最高,茉莉酸甲酯次之。LrPGIP启动子片段的长度为706 bp,具有激素以及胁迫响应等多个顺式作用元件。将LrPGIP启动子驱动β-葡萄糖苷酸酶(GUS)基因的表达框转入烟草表达,G US活性明显受尖孢镰刀菌、交链格孢侵染以及氯化汞、氯化钠胁迫的诱导,同时还响应茉莉酸、水杨酸等防卫相关植物激素。氯化汞处理后的GUS活性上调幅度最大,其次是交链格孢和乙烯利。可见,LrPGIP启动子活性受植物激素、生物及非生物胁迫的诱导。上述结果表明,LrPGIP可能是岷江百合抵御尖孢镰刀菌侵染的重要抗病基因。

关键词：岷江百合多聚半乳糖醛酸酶抑制蛋白表达特性启动子活性

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三七内切-β-1,4-葡聚糖酶基因PnCel1的表达及功能分析

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西北植物学报 2023年第5期43卷 732-741页

作者：苏琳琳王瀚林甘昆发陈晓华刘迪秋昆明理工大学生命科学与技术学院昆明650500 云南省三七资源可持续利用重点实验室昆明650500

内切-β-1,4-葡聚糖酶(endo-1,4-β-glucanases,EGases)广泛参与植物细胞壁的编辑,在组织伸长、果实成熟和脱落等过程中起重要作用。该研究采用RT-PCR方法,从三七(Panax notoginseng)中克隆了1个EGases基因(PnCel1),并对其进行表达和功... 详细信息

内切-β-1,4-葡聚糖酶(endo-1,4-β-glucanases,EGases)广泛参与植物细胞壁的编辑,在组织伸长、果实成熟和脱落等过程中起重要作用。该研究采用RT-PCR方法,从三七(Panax notoginseng)中克隆了1个EGases基因(PnCel1),并对其进行表达和功能分析。结果显示:(1)外源茉莉酸甲酯、水杨酸、赤霉素、脱落酸和乙烯利处理显著诱导PnCel1的表达,而根腐病菌茄腐镰刀菌(Fusarium solani)、尖孢镰刀菌(Fusarium oxysporum)以及交链格孢(Alternaria alternata)、木贼镰刀菌(Fusarium equiseti)侵染三七后显著抑制PnCel1的表达。(2)亚细胞定位分析表明,PnCel1-GFP融合蛋白定位于洋葱表皮细胞的细胞壁中。(3)采用染色体步移技术克隆出PnCel1的启动子序列[(-1)~(-828)bp],进而成功构建PnCel1启动子驱动的β-葡萄糖醛酸酶植物表达载体(pBI121-PPnCel1-GUS)并转入烟草(Nicotiana tabacum L.),经PCR筛选鉴定获得阳性转基因烟草7株。(4)GUS活性检测结果表明,5种植物激素能诱导PnCel1的启动子活性,但茄腐镰刀菌等4种病原菌侵染明显降低了PnCel1启动子的转录活性,且PnCel1启动子受三七WRKY转录因子PnWRKY5/9/12/15/27的负调控。(5)PnCel1过表达转基因烟草与野生型烟草相比,对茄腐镰刀菌的易感性增加,木质素含量降低,表明PnCel1可能参与改变细胞壁的结构。研究表明,植物激素能上调三七根中PnCel1的表达,而病原菌侵染降低了PnCel1的表达水平,并抑制PPnCel1的活性,推测三七PnCel1可能通过改变细胞壁结构而增加三七对根腐病菌的易感性。

关键词：三七茄腐镰刀菌内切-β-1,4-葡聚糖酶易感性

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Cloning of Porcine Lactoferrin Gene and Construction of Expression System in Recombinant Lactobacillus

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Journal of Northeast Agricultural University(English Edition) 2011年第4期18卷 32-38页

作者： ZONG Xiaolin HA Zhuo ZHAO Lili liu diqiu QIAO Xinyuan JIANG Yanping GE Junwei LI Yijing TANG Lijie College of Life Sciences Northeast Agricultural University Harbin 150030 China College of Veterinary Medicine Northeast Agricultural University Harbin 150030 China

Lactobacillus was selected as a bacterial carrier for expression of N-lobe of porcine lactoferrin （PLFN）. A pair of primers was designed with Oligo6.0 and used to amplify PLFN gene. It was in accordance with the characters of translational fusions from gene and expression vector plasmid. A 1 077 bp fragment of the gene from PLF was cloned from mammary gland tissue of the lactating sow on the third day by RT-PCR; the gene was connected with the vector plasmid pPG612.1 and transformed into the host strain JM109. The recombinant expression vector plasmid pPG612-PLFN was created and identified by using plasmid extraction, PCR, restriction enzyme digestion and sequence analysis. The recombinant plasmid was transformed into Lactobacillus casei ATCC393, Lactobacillus plantarum KLDS 1.0344, Lactobacillus paracasei KLDS 1.0652 and Lactobacillus pentosus KLDS 1.0413 by electroporation, and produced the recombinant strains of pPG612-PLFN/L, casei, pPG612-PLFN/L, plantarum, pPG612-PLFN/ L. paracasei and pPG612-PLFN/L, pentosus, respectively. The results indicated that PLFN gene had inserted into the expression vectors and achieved multiple Laetobacillus expression systems. It electes the base for the expression and production of recombinant porcine lactoferrin in Lactobaeillus

关键词： porcine lactoferrin Lactobacillus expression vector system

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