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【背景】小麦叶疫病菌于20世纪60年代入侵我国后,迅速传播扩散并在局部区域造成严重危害,对我国小麦的健康发展构成了巨大威胁。【方法】设计出检测小麦叶疫病菌的特异性引物,建立快速检测该病菌的PCR方法。用真菌通用引物ITS4/ITS5对小麦叶疫病菌进行PCR扩增,将扩增产物进行克隆和测序,使用DNAMAN软件设计出检测该病菌的特异性引物LJY1和LJY2,优化PCR反应体系。【结果】建立了该病菌的PCR检测方法,PCR反应体系:25 mmol·L-1Mg Cl22.5μL,10 mmol·L-1d NTP 1.0μL,10μmol·L-1引物各0.5μL,DNA模板8 ng,最佳退火温度57.6℃。【结论与意义】该方法可以准确地将小麦叶疫病菌与其他链格孢属的真菌区分开。本研究结果为小麦叶疫病的快速检测提供了依据,能够有效防止该病菌在小麦进出口贸易中传入我国。 相似文献
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小麦根腐病菌索氏平脐蠕孢SYBR Green I实时荧光定量PCR检测技术研究 总被引:3,自引:0,他引:3
由索氏平脐蠕孢Bipolaris sorokiniana引起的小麦根腐病,常和其他土传真菌病害混合发生,传统的症状鉴别方法很难区分,导致病害防控难度增加。为建立病菌实时荧光定量检测体系,根据ITS序列设计引物,筛选出1对特异性引物BS‐F/R,扩增片段大小为280bp。以菌丝DNA为标准品构建实时荧光定量标准曲线,并对其灵敏度、特异性、可重复性进行评价。结果表明,建立的实时荧光定量PCR检测方法速度快,灵敏度高,特异性强,重复性好。构建的荧光定量PCR标准曲线循环阈值与模板浓度呈良好的线性关系,溶解曲线的吸收峰单一,扩增效率良好。利用该定量检测体系,可以检测出田间小麦样品中52.8fg/μL的病菌DNA。 相似文献
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广东省柑橘炭疽病病原菌的形态与分子鉴定 总被引:2,自引:0,他引:2
炭疽病是柑橘的主要真菌性病害之一。2007年春,广东省德庆县名优柑橘品种贡柑炭疽病暴发流行。为了明确该县及广东省其他地区柑橘炭疽病菌的种类,为防治提供依据,对采集自广东省6个地区柑橘属10个栽培品种上的炭疽病样本进行病原菌分离,共获得柑橘炭疽病菌单孢菌株75株,对其中10株代表性的菌株进行了种类鉴定。通过培养性状和形态学特征观测、核糖体DNA(rDNA)内转录间区(ITS)序列分析、ITS区特异性引物PCR检测和系统发育关系比较等方面的研究,结果表明:10个柑橘炭疽病菌菌株均为盘长孢状刺盘孢Colletotrichum gloeosporioides,未发现国际上其他国家报道的严重危害柑橘花器和幼果部位的柑橘花后落果病病原菌——尖刺盘孢C.acutatum。 相似文献
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本研究测定了褐飞虱Nilaparvata lugens、白背飞虱Sogatella furcifera和灰飞虱Laodelphax striatellus的rDNA ITS1和ITS2的序列, 以探讨这3种稻飞虱的分子鉴定方法。3种飞虱的ITS1和ITS2侧翼区(18S, 5.8S和28S)序列相对稳定, 但ITS1和ITS2序列在3种飞虱中变异较大。 ITS1在所分析的438个位点中可变位点达294个, ITS2在分析的403个位点中可变位点为177个。根据3种飞虱rDNA的ITS1和ITS2序列设计了特异性引物, 应用特异性引物对样品进行了PCR扩增, 分析发现3种飞虱ITS1区的特异性引物扩增效果不理想, 而ITS2区的特异性引物可以稳定地扩增出明显的目的DNA条带. 因此, 采用ITS2区的特异性引物可以对3种飞虱进行快速的分子鉴定。 相似文献
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油茶白绢病原菌齐整小核菌分子检测的研究 总被引:1,自引:0,他引:1
目的:设计特异性引物建立油茶白绢病齐整小核菌的快速分子检测体系。方法:扩增齐整小核菌核糖体DNA ITS区并测定其序列,比较该序列与GenBank中近似种的ITS序列差异,设计了特异性引物BF1和BR2。结果:该引物可以从齐整小核菌中扩增得到约540bp特异性条带,而扩增其它近似或相关菌株时没有相应的特异性条带。在25μL PCR反应体系中,引物BF1和BR2检测灵敏度为1pg浓度DNA。结论:利用设计的BF1和BR2特异性引物结合PCR方法可快速的扩增出齐整小核菌DNA,检测灵敏度为1pg.但在生产实践中诊断油茶白绢病发病前组织中的齐整小核菌还需要进一步研究。 相似文献
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为筛选得到优良植物病害生防菌,对广西生姜Zingiber officinale种植区健康生姜根系和叶片中的共生真菌进行了组织分离,以生姜茎腐病菌群结腐霉Pythium myriotylum和香蕉枯萎病菌尖孢镰刀菌古巴专化型4号生理小种Fusarium oxysporum f. sp. cubense race 4为指示菌,通过平板对峙培养法和发酵液菌落直径法试验进行筛选评价,并结合形态学观察及ITS序列分析对筛选出的生防效果最好的共生真菌进行了鉴定。结果表明,从生姜植株共分离得到34株共生真菌,其中根系分离22株,叶片分离12株;对峙培养发现有6株共生真菌对生姜茎腐病菌和香蕉枯萎病菌均有抑制作用;其中菌株SBM-11拮抗作用最强,对生姜茎腐病菌抑制率达到93%,对香蕉枯萎病菌抑制率达到82%;SBM-11的发酵液对生姜茎腐病菌和香蕉枯萎病菌抑制率分别为82%、73%,与其他菌株发酵液抑制效果相比差异明显;结合形态和分子鉴定结果表明SBM-11菌株为绿色木霉Trichoderma viride,极具生防潜力。 相似文献
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Q Sun Y Zhang S Zhu H Cui H Tian D Yan G Huang Z Zhu D Wang X Li H Jiang H An W Xu 《Journal of virology》2012,86(18):10228-10229
Coxsackievirus A1 (CVA1) belongs to human enterovirus species C within the family Picornaviridae, order Picornavirales. Two Chinese CVA1 isolates, HT-THLH02F/XJ/CHN/2011 and KS-ZPH01F/XJ/CHN/2011, were isolated from stool specimens of two healthy children in the Xinjiang Uygur autonomous region of China. They were found to elicit cytopathic effects in a human rhabdomyosarcoma cell line, and complete genome sequences of these two CVA1 isolates revealed that natural intertypic recombination events occurred between CVA1 and CVA22. 相似文献
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Guo Yan Wang Hongjiang Xu Songtao Zhou Hangyu Zhou Chao Fu Shihong Cheng Mengli Li Fan Deng Yongqiang Li Xiaofeng Wang Huanyu Qin Cheng-Feng 《中国病毒学》2021,36(1):113-121
West Nile virus(WNV) is an important neurotropic flavivirus that is widely distributed globally. WNV strain XJ11129 was first isolated in Xinjiang, China, and its genetic and biological characteristics remain largely unknown. In this study,phylogenetic and sequence analyses revealed that XJ11129 belongs to lineage 1 a and shares high genetic identity with the highly pathogenic strain NY99. Then, the full-length genomic c DNA of XJ11129 was amplified and assembled using a modified Gibson assembly(GA) method. The virus(named r XJ11129) was successfully rescued in days following this method. Compared with other wild-type WNV isolates, r XJ11129 exhibited virulence indistinguishable from that of the NY99 strain in vivo. In summary, the genomic and virulence phenotypes of r XJ11129 were characterized in vivo and in vitro, and these data will improve the understanding of the spread and pathogenesis of this reemerging virus. 相似文献
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《Journal of Plant Interactions》2013,8(3):262-270
The interaction between sunflower plants showing a high level of quantitative resistance and five Plasmopara halstedii (the causal agent of downy mildew) isolates of several races were studied using five single zoosporangium isolates per pathogen isolate. Aggressiveness criteria were analyzed for 25 P. halstedii single zoosporangium isolates. Based on the reaction for the P. halstedii isolates to four sunflower hybrids H1–H4 varying only in their downy mildew resistance genes, there were differences in virulence spectrum in pathogen isolates. Analysis of five single zoosporangium isolates for P. halstedii isolates showed significant variability within pathogen isolate for all aggressiveness criteria but not for all pathogen isolates. The hypothesis explaining the interaction between P. halstedii and its host plant was discussed on the level of pathogenicity. 相似文献
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The coumarin phytoalexins ayapin and scopoletin accumulate in longitudinal stem sections of sunflower (Helianthus annuus L., Compositae) following inoculation with fungi both pathogenic (Alternaria helianthi) and nonpathogenic (Helminthosporium carbonum) to this plant. Both compounds were induced more rapidly, and they attained higher levels in tissue inoculated with the heterologous pathogen H. carbonum as compared with the sunflower pathogen A. helianthi. Similarly, scopoletin and ayapin accumulated to comparatively low concentrations following inoculation with a second sunflower pathogen, Phoma macdonaldii. Scopoletin was biosynthesized de novo following inoculation, although levels of its glucoside scopolin exceeded those of the aglucone in both infected and control tissues. Both scopoletin and scopolin were routinely detected in trace amounts in uninoculated tissue. In contrast, ayapin was not detected as a component of uninfected plants. When [14C]scopoletin was supplied to induced sunflower stem sections about 36% of the recovered radioactivity was in the form of ayapin. In vitro studies demonstrated that A. helianthi possessed the ability to rapidly degrade both scopoletin and ayapin, whereas H. carbonum was much less efficient in these traits. The differential degradation of these compounds by phytopathogenic fungi which do not attack sunflower is also discussed. 相似文献
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目的:分析我国新疆伤寒沙门菌分离株XJ19与全基因组测序的国际标准菌株CT18的蛋白表达差异,并推算基因差异。方法:运用二维蛋白电泳,对CT18和XJ19在体外培养基中的全菌蛋白进行分离,使用PDQuest软件找到其差异蛋白,进行质谱鉴定;对CT18差异蛋白编码基因设计引物,以XJ19DNA为模板进行PCR扩增,检测CT18差异蛋白编码基因在XJ19的存在情况。结果:菌株XJ19中存在53个特异蛋白点,鉴定出47个,但这些蛋白的编码基因在CT18中均存在,其中36个蛋白点在CT18的蛋白谱中不存在,11个蛋白在CT18中处于其他修饰状态;CT18中找到13个特异蛋白点,质谱鉴定出7个,其中6个所对应基因在XJ19中均能扩增出目的片段,但点C9蛋白的编码基因在XJ19中不存在。菌株XJ19中的多个差异蛋白参与磷酸戊糖途径的代谢及信号感应调控,此外超氧化物歧化酶、外膜蛋白OmpA呈现与CT18不同的修饰状态。结论:我们认为不同伤寒沙门菌分离株的遗传差异不仅仅是基因的有或无,还包括蛋白的不同表达和修饰所造成的不同调控机制和代谢的差异。 相似文献
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David J. Evans Gerald B. Pier Michael J. Coyne Jr Joanna B. Goldberg 《Molecular microbiology》1994,13(3):427-434
Summary
Strains of Pseudomonas aeruginosa initially isolated from patients with cystic fibrosis (CF) often express a smooth lipopolysaccharide (LPS) containing many long O side-chain antigens, but once a chronic infection is established, strains recovered from these patients express little or no LPS O antigen. The genetic basis for this loss of O antigen expression by P. aeruginosa CF isolates is unknown. We report here that 20 CF isoiates of P. aeruginosa , 13 of which are LPS-rough, were each capable of expressing serogroup 011 antigen when provided with the rfb iocus from P. aeruginosa serogroup 011 strain PA103 on the recombinant plasmid pLPS2. Eight of the thirteen LPS-rough isolates co-expressed another, presumably endogenous, O antigen when they contained pLPS2. Different subcloned regions of pLPS2 complemented distinct strains to restore endogenous O antigen expression. These data suggest that the loss of O antigen expression by P. aeruginosa CF isolates results from alterations specific to the rfb region, and is not due to mutations involving other loci or ancillary LPS genes. 相似文献
Strains of Pseudomonas aeruginosa initially isolated from patients with cystic fibrosis (CF) often express a smooth lipopolysaccharide (LPS) containing many long O side-chain antigens, but once a chronic infection is established, strains recovered from these patients express little or no LPS O antigen. The genetic basis for this loss of O antigen expression by P. aeruginosa CF isolates is unknown. We report here that 20 CF isoiates of P. aeruginosa , 13 of which are LPS-rough, were each capable of expressing serogroup 011 antigen when provided with the rfb iocus from P. aeruginosa serogroup 011 strain PA103 on the recombinant plasmid pLPS2. Eight of the thirteen LPS-rough isolates co-expressed another, presumably endogenous, O antigen when they contained pLPS2. Different subcloned regions of pLPS2 complemented distinct strains to restore endogenous O antigen expression. These data suggest that the loss of O antigen expression by P. aeruginosa CF isolates results from alterations specific to the rfb region, and is not due to mutations involving other loci or ancillary LPS genes. 相似文献
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Internal Transcribed Spacer Sequence Analysis of Puccinia helianthi Schw. and Its Application in Detection of Sunflower Rust
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Dandan Guo Jiali Jing Wenjie Hu Shrishail S. Navi Lan Jing 《Journal of Phytopathology》2016,164(2):141-146
Two basidiomycete‐specific primers ITS1‐F and ITS4‐B were used in identification of the genus Puccinia. The primers showed good specificity for the genus with an 816‐bp product that was amplified exclusively. Twenty sequences of internal transcribed spacer (ITS) regions of Puccinia helianthi isolates from China remain unchanged. The whole ITS length (including ITS1 sequence 194 bp, 5.8S rRNA gene 156 bp, ITS2 sequence 206 bp) was 556 bp. By comparing the aligned ITS sequences of several Puccinia isolates from China, Spain and the United States, ITS homogeneity among these sunflower rust isolates was >99%. Genetic homology and phylogeny of P. helianthi with other Puccinia spp. was investigated. Nineteen sequences of rDNA ITS1 and ITS2 were determined and used as phylogenetic markers. Phylogenetic analysis of ITS regions showed that Puccinia spp. of sunflower was clustered in one clade with P. komarovii and P. violae, divergent from Puccinia spp. of Chrysanthemum, P. tenaceti of tansy (Tanacetum vulgare) and Puccina spp. of big sagebrush (Artemisia tridentate) indicating sunflower rust had distant phylogenetic relationships with other Compositae rusts. With the specified primers SR‐1 and SR‐2, either from purified urediniospores or symptomless (but infected) sunflower leaves could be examined specifically. Therefore, results of this study help in detection and polygenetic study of rust fungi occurring on sunflower. 相似文献
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Nachaat Sakr 《Archives Of Phytopathology And Plant Protection》2013,46(14):1633-1641
Aggressiveness variation and its alternation with non-race specific resistance in sunflower were studied in 19 Plasmopara halstedii isolates belonging to several races. Regarding aggressiveness criteria, percentage infection, latent period, sporulation density and dwarfing, on two sunflower inbred lines showing different levels of non-race specific resistance resistance FU and BT, there were significant differences in aggressiveness for P. halstedii isolates. The index of aggressiveness varied between 9.4 and 31.4. The inbred line BT, rather susceptible in the field, showed a higher percentage infection, a higher sporulation density, a shorter latent period and less reduced hypocotyl length than inbred line FU, which showed a greater resistance in the field. Percentage infection on FU was 1.4% less than BT, latent period on BT was 12.4% less than FU, sporulation density on FU was 22.3% less than BT and reduced hypocotyl length on BT was 15.3% less than FU. Consequently, it seems that the criteria as latent period, sporulation density and reduction of hypocotyl length could be used to measure non-race specific resistance in sunflower to P. halstedii under controlled conditions. 相似文献
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Nachaat Sakr 《Archives Of Phytopathology And Plant Protection》2013,46(12):1386-1404
Morphological, pathogenic and genetic variation was studied in seven Plasmopara halstedii (sunflower downy mildew) isolates of several races using five singlezoosporangium isolates per pathogen isolate. Aggressiveness criteria were analysed in one sunflower inbred line showing a high level of quantitative resistance. Genetic relationships were detected between the single zoosporangium isolates using 12 expressed sequence tags (EST)-derived markers. Analysis of the five single zoosporangium isolates for P. halstedii isolates showed variability within pathogen isolates for all aggressiveness criteria, but not for all pathogen isolates. Isolates of races 100 and 3xx were characterised with shorter latent period and higher sporulation density than the isolate of races 7xx. All pathogen isolates showed high percentage infection values and caused a large reduction in seedling size except for one isolate involved in dwarfing. There was no relation between zoosporangia form or size and race virulence profiles or aggressiveness criteria. There was no intra-genetic variation for all pathogen isolates, but it was observed an important genetic variation between single zoosporangium isolates of all races. No correlation was detected between pathogenicity traits and EST genotypes. 相似文献