本文主要研究内容
作者戴蓬博（2019）在《粉红聚端孢引致富士苹果心腐病的侵染特征及致病机制》一文中研究指出：苹果心腐病是一种苹果主要果实病害。可造成采前落果、果实心室腐烂和储藏期烂果等,严重影响苹果品质、降低苹果产量,给苹果种植者、经营者和消费者均带来较大的经济损失。在我国,粉红聚端孢是引起苹果心腐病的主要病原。心腐病主要发生于苹果心室开放品种。红富士属于心室半开放品种,在果实发育早期,心室封闭,随着果实膨大心室腔逐渐开放。在果实发育过程中,病原菌的侵染时间、侵染部位都不清楚,严重影响了针对性防治策略的制定。本论文拟研究我国心腐病的主要病原菌粉红聚端孢在主栽富士品种的侵染特征和致病机制,以期为苹果心腐病的防治及粉红聚端孢的致病机理提供理论依据。取得以下主要研究结果:1.苹果心腐病为果实心室病害,果实的特殊结构造成病原菌侵入过程研究的困难性,本研究首先构建了PEG介导的粉红聚端孢遗传转化体系和绿色荧光标记菌株:通过液体培养基摇培粉红聚端孢获取幼嫩菌丝,用崩溃酶和裂解酶酶解幼嫩菌丝细胞壁制备原生质体,用PEG介导原生质体转化将携带博来霉素筛选标记和绿色荧光基因的质粒插入粉红聚端孢基因组获取有筛选抗性的转化子,通过筛选获得粉红聚端孢荧光标记菌株TR45。与野生型相比,该菌株菌落生长、产孢量和致病力均未发生明显改变,在不含筛选标记的培养皿中继代培养10代荧光仍能稳定表达,接种苹果组织后能观察到荧光亮度明显的侵染菌丝。可以作为进一步研究其基因功能及侵染特性的有效工具。2.借助荧光标记菌株TR45,进行了粉红聚端孢对苹果不同花组织、萼筒、萼心间组织和心室内部的侵染和扩展特征研究,结果发现:粉红聚端孢可以侵染苹果花瓣、花药、花丝、柱头和离生花柱等花器组织,并在花器残体上繁殖产生菌丝和分生孢子。在花落后2周左右,随着萼片逐渐闭合,携带菌丝和孢子的花器残体被包裹到萼筒内。在花后3周可以观察到粉红聚端孢从合生花柱顶缝进入合生花柱,在花柱缝隙组织内扩展;进而在萼心间组织的延伸缝中扩展;在花后60天菌丝扩展至心室腔间组织;在花后100天随着萼窦逐渐扩大,萼窦、萼心间延伸缝和心室腔连通,菌丝进入心室腔;在心室腔内,菌丝在心室壁上扩展,从心室壁裂缝进入果肉组织。另外还发现,从花后60天以后到采收前,粉红聚端孢分生孢子在萼筒内不能萌发,推测其花后2个月不能再侵入花柱。这些结果表明:合生花柱顶部缝隙为粉红聚端孢侵入苹果主要关口,而要成功侵入苹果果肉组织还需要经过花柱缝隙、萼心间组织延伸缝、心室壁裂缝等通道,以及萼窦的开放。3.除了侵染花器,粉红聚端孢侵染苹果萼洼处,在萼洼处形成的繁殖体也是心腐病潜在菌源。对粉红聚端孢在苹果表面侵染引起黑点病侵染特征研究发现:在蒸馏水中,12 h分生孢子萌发率仅为2%,苹果汁溶液中萌发率提高到99%,在气孔、皮孔、微裂痕处的萌发率显著高于周围苹果表皮,表明苹果汁成分可以显著促进粉红聚端孢孢子萌发。在相对湿度95%条件下18 h分生孢子萌发达到100%,显示粉红聚端孢侵染对湿度需求较低。粉红聚端孢不能直接穿透侵染苹果表皮,菌丝可以从气孔、皮孔、微裂痕等孔口处侵染。这些结果表明,果袋尤其是透气性差的膜袋所提供的微环境湿度使粉红聚端孢在苹果表面完成侵染循环比较容易满足,可以部分解释套袋后容易发生苹果黑点病的原因。黑点病主要发生在苹果萼洼处,黑点病的发生不但直接影响果实外观品质,而且可以为心腐病的发生提供菌源。4.粉红聚端孢在侵染苹果果皮和果肉的组织时,菌丝仅在真皮细胞和薄壁细胞的细胞间隙扩展,显示粉红聚端孢具有孢间扩展特性,不能穿透寄主细胞壁;进一步观察发现,在花柱缝隙扩展过程中,菌丝不突破花柱缝的内表皮细胞,在萼心间延伸缝扩展,菌丝不突破延伸缝的细胞内壁,在心室腔中,菌丝只能从心室壁上的裂缝进入果肉细胞,支持粉红聚端孢不具备穿透寄主细胞壁的能力。这可能是粉红聚端孢在侵染过程中不直接造成果实腐烂,仅在进入果实心室内部后才导致发病,引致心腐病发生的主要原因。5.对粉红聚端孢产生单端孢菌素(Tcin)的致病性作用及功能进行了研究,敲除粉红聚端孢合成通路的关键基因,单端孢烯合成酶基因(Tri5)并观察外源添加Tcin对致病的影响,发现不产Tcin的粉红聚端孢ΔTrTri5突变体在果实上完全失去致病力,互补Tri5基因使突变体单端孢菌素的合成能力回复后致病力也恢复,外源添加Tcin可以回复ΔTrTri5突变体的致病力;用少量Tcin(10μg/mL)处理苹果,此时果皮和果肉组织无明显症状,但与Tcin缺失突变体共同接种可以促进致病,说明单端孢菌素在粉红聚端孢的致病过程中起重要作用;用100μg/mL和500μg/mL单端孢菌素处理苹果后在苹果上不引起症状变化,但电镜观察发现,果肉细胞的质体内膜、外膜破裂、质体降解,细胞膜和液泡膜上有高密度电子密集物积累。这些结果显示,单端孢菌素具有植物毒素功能,单端孢菌素是粉红聚端孢致病过程中的关键致病因子。本研究发现合生花柱顶端的自然孔口花柱顶缝是粉红聚端孢侵入苹果的关键部位,病菌入侵后沿花柱裂缝、延伸缝、萼窦、心室腔裂缝最后扩展果肉,导致心室果肉腐烂。明确单端孢菌素是植物毒素,为粉红聚端孢侵染苹果引致心腐病的关键致病因子。该研究对于靶向新农药设计、病原与寄主互作机制研究,及针对性病害防治策略制定具有理论及实践理论意义。
Abstract
ping guo xin fu bing shi yi chong ping guo zhu yao guo shi bing hai 。ke zao cheng cai qian la guo 、guo shi xin shi fu lan he chu cang ji lan guo deng ,yan chong ying xiang ping guo pin zhi 、jiang di ping guo chan liang ,gei ping guo chong zhi zhe 、jing ying zhe he xiao fei zhe jun dai lai jiao da de jing ji sun shi 。zai wo guo ,fen gong ju duan bao shi yin qi ping guo xin fu bing de zhu yao bing yuan 。xin fu bing zhu yao fa sheng yu ping guo xin shi kai fang pin chong 。gong fu shi shu yu xin shi ban kai fang pin chong ,zai guo shi fa yo zao ji ,xin shi feng bi ,sui zhao guo shi peng da xin shi qiang zhu jian kai fang 。zai guo shi fa yo guo cheng zhong ,bing yuan jun de qin ran shi jian 、qin ran bu wei dou bu qing chu ,yan chong ying xiang le zhen dui xing fang zhi ce lve de zhi ding 。ben lun wen ni yan jiu wo guo xin fu bing de zhu yao bing yuan jun fen gong ju duan bao zai zhu zai fu shi pin chong de qin ran te zheng he zhi bing ji zhi ,yi ji wei ping guo xin fu bing de fang zhi ji fen gong ju duan bao de zhi bing ji li di gong li lun yi ju 。qu de yi xia zhu yao yan jiu jie guo :1.ping guo xin fu bing wei guo shi xin shi bing hai ,guo shi de te shu jie gou zao cheng bing yuan jun qin ru guo cheng yan jiu de kun nan xing ,ben yan jiu shou xian gou jian le PEGjie dao de fen gong ju duan bao wei chuan zhuai hua ti ji he lu se ying guang biao ji jun zhu :tong guo ye ti pei yang ji yao pei fen gong ju duan bao huo qu you nen jun si ,yong beng hui mei he lie jie mei mei jie you nen jun si xi bao bi zhi bei yuan sheng zhi ti ,yong PEGjie dao yuan sheng zhi ti zhuai hua jiang xie dai bo lai mei su shai shua biao ji he lu se ying guang ji yin de zhi li cha ru fen gong ju duan bao ji yin zu huo qu you shai shua kang xing de zhuai hua zi ,tong guo shai shua huo de fen gong ju duan bao ying guang biao ji jun zhu TR45。yu ye sheng xing xiang bi ,gai jun zhu jun la sheng chang 、chan bao liang he zhi bing li jun wei fa sheng ming xian gai bian ,zai bu han shai shua biao ji de pei yang min zhong ji dai pei yang 10dai ying guang reng neng wen ding biao da ,jie chong ping guo zu zhi hou neng guan cha dao ying guang liang du ming xian de qin ran jun si 。ke yi zuo wei jin yi bu yan jiu ji ji yin gong neng ji qin ran te xing de you xiao gong ju 。2.jie zhu ying guang biao ji jun zhu TR45,jin hang le fen gong ju duan bao dui ping guo bu tong hua zu zhi 、e tong 、e xin jian zu zhi he xin shi nei bu de qin ran he kuo zhan te zheng yan jiu ,jie guo fa xian :fen gong ju duan bao ke yi qin ran ping guo hua ban 、hua yao 、hua si 、zhu tou he li sheng hua zhu deng hua qi zu zhi ,bing zai hua qi can ti shang fan shi chan sheng jun si he fen sheng bao zi 。zai hua la hou 2zhou zuo you ,sui zhao e pian zhu jian bi ge ,xie dai jun si he bao zi de hua qi can ti bei bao guo dao e tong nei 。zai hua hou 3zhou ke yi guan cha dao fen gong ju duan bao cong ge sheng hua zhu ding feng jin ru ge sheng hua zhu ,zai hua zhu feng xi zu zhi nei kuo zhan ;jin er zai e xin jian zu zhi de yan shen feng zhong kuo zhan ;zai hua hou 60tian jun si kuo zhan zhi xin shi qiang jian zu zhi ;zai hua hou 100tian sui zhao e dou zhu jian kuo da ,e dou 、e xin jian yan shen feng he xin shi qiang lian tong ,jun si jin ru xin shi qiang ;zai xin shi qiang nei ,jun si zai xin shi bi shang kuo zhan ,cong xin shi bi lie feng jin ru guo rou zu zhi 。ling wai hai fa xian ,cong hua hou 60tian yi hou dao cai shou qian ,fen gong ju duan bao fen sheng bao zi zai e tong nei bu neng meng fa ,tui ce ji hua hou 2ge yue bu neng zai qin ru hua zhu 。zhe xie jie guo biao ming :ge sheng hua zhu ding bu feng xi wei fen gong ju duan bao qin ru ping guo zhu yao guan kou ,er yao cheng gong qin ru ping guo guo rou zu zhi hai xu yao jing guo hua zhu feng xi 、e xin jian zu zhi yan shen feng 、xin shi bi lie feng deng tong dao ,yi ji e dou de kai fang 。3.chu le qin ran hua qi ,fen gong ju duan bao qin ran ping guo e wa chu ,zai e wa chu xing cheng de fan shi ti ye shi xin fu bing qian zai jun yuan 。dui fen gong ju duan bao zai ping guo biao mian qin ran yin qi hei dian bing qin ran te zheng yan jiu fa xian :zai zheng liu shui zhong ,12 hfen sheng bao zi meng fa lv jin wei 2%,ping guo zhi rong ye zhong meng fa lv di gao dao 99%,zai qi kong 、pi kong 、wei lie hen chu de meng fa lv xian zhe gao yu zhou wei ping guo biao pi ,biao ming ping guo zhi cheng fen ke yi xian zhe cu jin fen gong ju duan bao bao zi meng fa 。zai xiang dui shi du 95%tiao jian xia 18 hfen sheng bao zi meng fa da dao 100%,xian shi fen gong ju duan bao qin ran dui shi du xu qiu jiao di 。fen gong ju duan bao bu neng zhi jie chuan tou qin ran ping guo biao pi ,jun si ke yi cong qi kong 、pi kong 、wei lie hen deng kong kou chu qin ran 。zhe xie jie guo biao ming ,guo dai you ji shi tou qi xing cha de mo dai suo di gong de wei huan jing shi du shi fen gong ju duan bao zai ping guo biao mian wan cheng qin ran xun huan bi jiao rong yi man zu ,ke yi bu fen jie shi tao dai hou rong yi fa sheng ping guo hei dian bing de yuan yin 。hei dian bing zhu yao fa sheng zai ping guo e wa chu ,hei dian bing de fa sheng bu dan zhi jie ying xiang guo shi wai guan pin zhi ,er ju ke yi wei xin fu bing de fa sheng di gong jun yuan 。4.fen gong ju duan bao zai qin ran ping guo guo pi he guo rou de zu zhi shi ,jun si jin zai zhen pi xi bao he bao bi xi bao de xi bao jian xi kuo zhan ,xian shi fen gong ju duan bao ju you bao jian kuo zhan te xing ,bu neng chuan tou ji zhu xi bao bi ;jin yi bu guan cha fa xian ,zai hua zhu feng xi kuo zhan guo cheng zhong ,jun si bu tu po hua zhu feng de nei biao pi xi bao ,zai e xin jian yan shen feng kuo zhan ,jun si bu tu po yan shen feng de xi bao nei bi ,zai xin shi qiang zhong ,jun si zhi neng cong xin shi bi shang de lie feng jin ru guo rou xi bao ,zhi chi fen gong ju duan bao bu ju bei chuan tou ji zhu xi bao bi de neng li 。zhe ke neng shi fen gong ju duan bao zai qin ran guo cheng zhong bu zhi jie zao cheng guo shi fu lan ,jin zai jin ru guo shi xin shi nei bu hou cai dao zhi fa bing ,yin zhi xin fu bing fa sheng de zhu yao yuan yin 。5.dui fen gong ju duan bao chan sheng chan duan bao jun su (Tcin)de zhi bing xing zuo yong ji gong neng jin hang le yan jiu ,qiao chu fen gong ju duan bao ge cheng tong lu de guan jian ji yin ,chan duan bao xi ge cheng mei ji yin (Tri5)bing guan cha wai yuan tian jia Tcindui zhi bing de ying xiang ,fa xian bu chan Tcinde fen gong ju duan bao ΔTrTri5tu bian ti zai guo shi shang wan quan shi qu zhi bing li ,hu bu Tri5ji yin shi tu bian ti chan duan bao jun su de ge cheng neng li hui fu hou zhi bing li ye hui fu ,wai yuan tian jia Tcinke yi hui fu ΔTrTri5tu bian ti de zhi bing li ;yong shao liang Tcin(10μg/mL)chu li ping guo ,ci shi guo pi he guo rou zu zhi mo ming xian zheng zhuang ,dan yu Tcinque shi tu bian ti gong tong jie chong ke yi cu jin zhi bing ,shui ming chan duan bao jun su zai fen gong ju duan bao de zhi bing guo cheng zhong qi chong yao zuo yong ;yong 100μg/mLhe 500μg/mLchan duan bao jun su chu li ping guo hou zai ping guo shang bu yin qi zheng zhuang bian hua ,dan dian jing guan cha fa xian ,guo rou xi bao de zhi ti nei mo 、wai mo po lie 、zhi ti jiang jie ,xi bao mo he ye pao mo shang you gao mi du dian zi mi ji wu ji lei 。zhe xie jie guo xian shi ,chan duan bao jun su ju you zhi wu du su gong neng ,chan duan bao jun su shi fen gong ju duan bao zhi bing guo cheng zhong de guan jian zhi bing yin zi 。ben yan jiu fa xian ge sheng hua zhu ding duan de zi ran kong kou hua zhu ding feng shi fen gong ju duan bao qin ru ping guo de guan jian bu wei ,bing jun ru qin hou yan hua zhu lie feng 、yan shen feng 、e dou 、xin shi qiang lie feng zui hou kuo zhan guo rou ,dao zhi xin shi guo rou fu lan 。ming que chan duan bao jun su shi zhi wu du su ,wei fen gong ju duan bao qin ran ping guo yin zhi xin fu bing de guan jian zhi bing yin zi 。gai yan jiu dui yu ba xiang xin nong yao she ji 、bing yuan yu ji zhu hu zuo ji zhi yan jiu ,ji zhen dui xing bing hai fang zhi ce lve zhi ding ju you li lun ji shi jian li lun yi yi 。
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论文作者分别是来自西北农林科技大学的戴蓬博，发表于刊物西北农林科技大学2019-07-11论文，是一篇关于粉红聚端孢论文,合生花柱论文,自然孔口论文,单端孢菌素论文,致病性论文,黑点病论文,西北农林科技大学2019-07-11论文的文章。本文可供学术参考使用，各位学者可以免费参考阅读下载，文章观点不代表本站观点，资料来自西北农林科技大学2019-07-11论文网站，若本站收录的文献无意侵犯了您的著作版权，请联系我们删除。
标签：粉红聚端孢论文; 合生花柱论文; 自然孔口论文; 单端孢菌素论文; 致病性论文; 黑点病论文; 西北农林科技大学2019-07-11论文;