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molecular detection of Phytophthora colocasiae of Taro Leaf Blight Based on PCR

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Journal of Landscape Research 2020年第1期12卷 33-35,38页

作者： HUANG Weihua YAN Meixin Biotechnology Research Institute Guangxi Academy of Agricultural Sciences

The present PCR assay was conducted to develop rapid and sensitive detection of Phytophthora colocasiae,in order to provide a robust and reliable tool for healthy seedling production of taro and limiting the transmission and spread of the causal organism of taro leaf blight in taro planting regions.The samples were used to extract total DNA and to be detected by PCR with P.colocasiae specific primer pairs PCSP-RL F/PCSP-RL R and PCSP-T F/PCSP-T R,respectively.Distinct fragments of about 200 bp and 240 bp were amplified by PCR using primers PCSP-RL F/PCSP-RL R and PCSP-T F/PCSP-T R,respectively.The analysis of the nucleotide sequence of the PCR products were found to be 99% identical to sequence of RAS-related protein (Ypt1) and phospho-ribosylanthranilate isomerase (TRP1) in P.colocasiae,respectively.It is concluded that rapid and sensitive developed PCR assay for detection of P.colocasiae could be used in routine diagnosis and aid in management practices to mitigate taro leaf blight.

关键词： Colocasia esculenta(L.)Schott Taro leaf blight Phytophthora colocasiae molecular detection

molecular detection and Disease Resistance Identification of Transgenic Wheat with pti5-vp16 Gene

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Agricultural Sciences in China 2003年第9期2卷 982-986页

作者： WEISong-hong CAOYuan-yin ZHANG-Yan-zhen ZHANGLing-bing WANGGang YANGJia-shu InstituteofPlantImmunology ShenyangAgriculturalUniversityShenyang110161P.R.China ResearchCenterforPlantGeneticEngineering theQuartermasterUniversityofChinesePeople‘sLiberationArmyChangchun130062P.R.China

The immature embryos of wheat cultivars Liaochun10, Tiechun1 and Fengqiang3 were bombarded with gold particles coated with pti5 vp16 by gene gun and disease resistant regenerated plants were attained. In order to confirm that the plants are genuine transformed ones, a series of molecular tests were conducted as follows. Firstly, transient GUS expression test on embryos two days after bombardment was done. There were many obvious blue spots produced on the surface of bombarded embryos after GUS staining, in which the maximum reached to 85 blue spots per embryo. Secondly, PCR test was performed with DNA from the regenerated plants obtained after double selection with ppt. 6 plants were found PCR test positive. At last, further verification analysis using dot hybridization and southern blotting was carried out on those PCR positive plants and the strong hybridization signals appeared as expected. All the above tests were uniformly indicated that the disease resistant regenerated plants were true transgenic plants. When inoculated with Blumeria graminis, transgenic wheat plants of PCR positive results were mostly resistant(R) after 7 days, and resis tant, moderate resistant(MR), moderate susceptible(MS) at 14 days respectively. The disease severity of them was distinctively lighter than that of control.

关键词： Transgenic wheat pti5 vp16 gene molecular detection Disease resistance

molecular detection of Main Diseases of Introduced Sugarcane in Guangxi

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Agricultural Biotechnology 2015年第1期4卷 33-35页

作者： Jinju WEI Zhanyun DENG Weihua HUANG Zhenqiang QIN Yangrui LI Hong HE Hongqin TANG Qiang LI Wenzan WANG Sugarcane Research Institute Guangxi Academy of Agricultural Sciences Sugarcane Research Center Chinese Academy of Agricultural Sciences Guangxi Key Laboratory for Sugarcane Biotechnology and Genetic Improvement Ministry of Agriculture Guangxi University

According to the quarantine procedures for sugarcane introduction, thirty-three sugarcane varieties/materials from the United States, Bangladesh, Thailand and some other countries were quarantined. The result showed that main diseases and pests that are concerned in quarantine were undiscovered. In addition, molecular detection results of Fiji disease and leaf scald disease were negative ; detection results of mosaic in six varleties/materials were positive; detection results of yellow leaf syndrome in eight varieties/materials were positive.

关键词： Introduction of sugarcane varieties Main diseases Quarantine molecular detection

molecular Techniques for detection of Major Diseases in Sugarcane

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Agricultural Biotechnology 2017年第2期6卷 1-5页

作者： Wenfeng LI Chunhua PU Yingkun HUANG Xiaoyan WANG Hongli SHAN Rongyue ZHANG Jiong YIN Zhiming LUO Yunnan Provincial Key Laboratory of Sugarcane Genetic Improvement Sugarcane Research Institute Yunnan Academy of Agricultural Sciences Kaiyuan 661699 China Yunnan Academy of Agricultural Sciences Kunming 650205 China

It is necessary to rapidly diagnose diseases, identify and monitor the pathogens to achieve scientific and effective control of major diseases in sugarcane. In the present study, the molecular techniques for rapid detection of 13 pathogens that cause 10 important diseases of sugarcane including smut, rust, leaf scald, ratoon stunting, red stripe, mosaic, Fiji, yellow leaf, white leaf and bacilliform virus were established by Sugarcane Research Institute, Yunnan Academy of Agricul- tural Sciences after years of effort. The results will provide scientific basis for effective diagnosis and control of sugarcane diseases, detection of virus-free seedlings and quarantine management of exotic species.

关键词： Sugarcane Key diseases molecular detection

molecular detection and quantification of Xanthomonas albilineans in juice from symptomless sugarcane stalks using a real-time quantitative PCR assay

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Molecular detection and quantification of Xanthomonas albili...

中国植物病理学会2021年学术年会

作者： SHI Yang ZHAO Jianying ZHOU Jingru NTAMBO Mbuya Sylvain XU Pengyuan ROTT Philippe C. GAO Sanji National Engineering Research Center for Sugarcane Fujian Agriculture and Forestry University Département de phytotechnie Faculté des Sciences AgronomiquesUniversité de Lubumbashi CIRAD UMRPHIMF-34398 MontpellierFranceand PHIMPlant Health InstituteUniv MontpellierCIRADINRAEInstitut AgroIRD

Leaf scald,a bacterial disease caused by Xanthomonas albilineans(Ashby) Dowson,is a major limiting factor for sugarcane production worldwide.Accurate identification and quantification of X.albilineans is a prerequisite for successful management of this disease.A very sensitive and robust qPCR assay was developed in this study for detection and quantification of X.albilineans using TaqMan probe and primers targeting a putative adenosine triphosphate-binding cassette(ABC) transporter gene(abc).The novel qPCR assay was highly specific to the 43 tested X.albilineans strains belonging to different pulsed-field gel electrophoresis(PFGE) groups.The detection thresholds were 100 copies/μL of plasmid DNA,100 fg/μL of bacterial genomic DNA,and 100 CFU/mL of bacterial suspension prepared from pure culture.This qPCR assay was 100 times more sensitive than a conventional PCR assay.The pathogen was detected by qPCR in 75.1%(410/546) symptomless stalk samples,whereas only28.4%(155/546) samples tested positive by conventional PCR.Based on qPCR data,population densities of X.albilineans in symptomless stalks of the same varieties differed between two sugarcane production areas in China,Beihai(Guangxi province) and Zhanjiang(Guangdong province).Nonsignificant correlation of X.albilineans populations was found between Beihai and Zhanjiang.The newly developed qPCR assay proved to be highly sensitive and reliable for the detection and quantification of X.albilineans in sugarcane stalks.

关键词： Leaf scald molecular detection real-time quantitative PCR Saccharum spp. Xanthomonas albilineans

molecular detection of Potato virus X（PVX）,Potato virus Y（PVY）,Potato leaf roll virus（PLRV）and Potato virus S（PVS）from Bangladesh

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Molecular detection of Potato virus X（PVX）,Potato virus Y...

中国植物病理学会2017年学术年会

作者： M.Rashid Zhang Xiaoyang Wang Ying Han Chenggui Department of Plant Pathology and State Key Laboratory for Agro-Biotechnology China Agricultural University

Potato（Solanum tuberosum） is considered as the fourth most important staple food source after rice（Oryza sativa）,maize（Zea mays） and wheat（Triticum aestivum） which is the most popular and important vegetable in Bangladesh.But it is frequently affected by different viruses and cause a huge yield loses.Viruses known to infect potato in Bangladesh include Potato virus X（PVX）,Potato virus Y（PVY）,Potato leaf roll virus（PLRV） and Potato virus S（PVS）.Though the increasing incidence of PVX,PVY and PLRV in main potato growing areas of Bangladesh is getting an alarming position andcaused significant yield losses,no molecular study had done before this work.In this research,molecular detection of potato viruses was done from the potato tuber collected from Munshiganj and Jessore districts of Bangladesh.To detect the plant viruses,total RNAs were extracted from the virus affected potato tubers and complementary cDNAs were synthesized.Reverse transcriptase polymerase chain reaction（RT-PCR） based detection conditions were optimized by using coat protein（CP） gene specific primers.After RT-PCR we found the amplified fragments of expected length for PVX,PVY,PLRV and PVS that were confirmed by subsequent sequencing.This is the first report for PVS in Bangladesh.Our resuits will be helpful for further molecular study on PVX,PVY,PLRV and PVS and their management.

关键词： molecular detection Potato virus X（PVX） Potato virus Y（PVY） Potato leaf roll virus（PLRV） Potato virus S（PVS） Bangladesh

molecular detection and Sequence Analysis of Polygalacturonase Szpg6 to Szpg10 Gene of Zebra Disease of Sisal

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Molecular Detection and Sequence Analysis of Polygalacturona...

中国植物病理学会2016年学术年会

作者： WU Wei-huai ZHENG Jin-long XI Jin-gen ZHENG Xiao-lan LIANG Yan-qiong LI Rui HE Chun-ping YI Ke-xian Environment and Plant Protection Institute CATAS/Hainan Key Laboratory for Detection and Control of Tropical Agricultural Pests

Sisal zebra disease caused by Phytophora nicotianae var.parasitica is a kind of main diseases which can cause serious damage to the sisal.In the present study,five specific primer pairs were designed in coding region based on the reference sequence of pppg6 to pppg10 gene from Phytophora parasitic.Five specific primer pairs were used for the molecular detection and gene cloning from sisal zebra pathgoen DNA.The results showed that Szpg6（sisal zebra polygalacturonase 6） to Szpg10 genes were conserved in all the tested sisal zebra isolates.Sequence alignment with the reference sequences of pppg6 to pppg10 gene from Phytophora parasitic revealed that 1,3,1,6 and 4 SNPs were identified in the Szpg6 to Szpg10 genes coding regions,respectively.There,five and three of these mutations were nonsynonymous in Szpg7,Szpg9 and Szpg10,and the rest were synonymous.Thus we speculated that there were some differences in function when Szpg7,Szpg9 and Szpg10 genes were compared to pppg7,pppg9 and pppg10.The results have laid a solid foundation for the further study of the role of sisal zebra germs polygalaturonase in the pathogenic process.

关键词： Zebra disease of sisal Polygalaturonase molecular detection

PCR-Mediated molecular detection of Hemileia vastatrix in Coffee

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PCR-Mediated Molecular Detection of Hemileia vastatrix in Co...

中国植物病理学会2017年学术年会

作者：吴伟怀汪涵 Li Le He Chunping Liang Yanqiong Zheng Jinlong Li Rui Huang Xin 易克贤 Environment and Plant Protection Institute CATAS/Ministry of Agriculture Key Laboratory for Monitoring and Control of Tropical Agricultural and Forest Invasive Alien PestsHainan Key Laboratory for Detection and Control of Tropical Agricultural Pests Institute of Tropical Agriculture and Forestry Hainan University

Coffee leaf rust is the most important fungal disease affecting coffee production,which worldwide caused by the biotrophic pathogen Hemileia vastatrix（Hv）.Thus,the rapid,accurate and identification of the pathogen is urgently required to manage and prevent the spreading of coffee leaf rust.In order to establish a molecular biological way for rapid test that has the characteristics of high specificity,high sensitivity and simple operation.This study were analysised by multiple comparisons among coffee rust rDNA-ITS and its proximal sequences,then gets the specific rDNA-ITS regions of Hv that was designed a pair of primers,Hv-ITS-F/R.The specificity of PCR assay were evaluated on the genome DNA from Hv,coffee brown spot pathogen,hyperparasite of Hv,coffee Colletotrichum and other fungi belonging to different Genus.The result showed that they only amplified 396 bp specific bands from Hv,not from other close or similar pathogenic fungi.The sensitivity of PCR assay was also evaluated.The detection sensitivity of the nested PCR was approximately 10 pg/μL at the DNA level.Via a further test of healthy and diseased coffee plants,the examining method has a great application prospect.These results illustrated that the primers Hv-ITS-F/R can be used to detect Hv in coffee tissue.Therefore,this method will be very useful in forecast of coffee rust early disease,growth and disease epidemiology.

关键词： Coffee Hemileia vastatrix molecular detection

Development of a loop‑mediated isothermal amplification assay for detection of Austropeplea tomentosa from environmental water samples

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Animal Diseases 2023年第1期3卷 35-48页

作者： Lily Tran Vignesh A.Rathinasamy Travis Beddoe Department of Animal Plant and Soil SciencesSchool of AgricultureBiomedicine and EnvironmentLa Trobe UniversityBundooraVIC 3083Australia Full list of author information is available at the end of the article

Lymnaeid snails are key intermediate hosts for the development and survival of Fasciola spp.,the causative agent of Fascioliasis which are economically important parasites infecting humans and livestock globally.The current control method for treating Fascioliasis is heavily reliant on anthelmintic drugs,particularly Triclabendazole(TCBZ)which has resulted in drug-resistant parasites and poses significant risk as there are no long-term efficacious alternatives available.Sustainable control measures at the farm level could include both parasite and snail control will play an important role in Fasciola spp.control and reduce the reliance on anthelmintic drugs.Implementation of such sustainable control measures requires effective identification of snails on the property however Lymnaeid snails are small and difficult to physically locate.Snail identification using an environmental DNA approach is a recent approach in which physically locating snails are not required.Austropeplea tomentosa,is the primary intermediate snail host for F.hepatica transmission in South-East Australia and we present an in-field loop-mediated isothermal amplification and water filtering method for the detection of A.tomentosa eDNA from water samples to improve current surveillance methods.This methodology is highly sensitive with a detection limit of 5×10^(−6)ng/μL,detected in<20 minutes,with cumulative sample preparation and amplification time under 1 hour.This proposed workflow could assist in monitoring areas to determine the risk of Fascioliasis infection and implement strategies to manage snail populations to ultimately reduce the risk of infection for humans and livestock.

关键词： Fasciola spp. Snail molecular detection DNA diagnostics LAMP Environmental sampling eDNA