• Title/Summary/Keyword: internal transcribed spacer(ITS)

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A Loop-mediated Isothermal Amplification Method for White-backed Planthopper-specific Detection (고리매개등온증폭법(LAMP)을 이용한 흰등멸구 특이 판별법)

  • Seo, Bo Yoon;Park, Chang Gyu;Jung, Jin Kyo;Cho, Jumrae;Lee, Gwan-Seok;Kim, Kwang-Ho
    • Korean journal of applied entomology
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    • v.57 no.4
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    • pp.393-399
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    • 2018
  • A loop-mediated isothermal amplification (LAMP) primer set (WBPH-65) was designed for the species-specific detection of white-backed planthopper (WBPH) Sogatella furcifera based on the full-length sequence of the internal transcribed spacer 2 (ITS2) (KC417469.1). The WBPH-65 primer set consists of six primers (total 165 bp), F3 (18 bp), B3 (18 bp), FIP (43 bp), BIP (40 bp), LF (21 bp), and LB (25 bp). After the LAMP reaction of three rice planthoppers, S. furcifera, Nilaparvata lugens, and Laodelphax striatellus, with the WBPH-65 primer set for 60 min at $65^{\circ}C$, the LAMP products were observed in the genomic DNA of S. furcifera only. According to the DNA amount of S. furcifera and incubation duration at $65^{\circ}C$, the difference of fluorescence relative to the negative control (0 ng) was clearly observed in a 40-min incubation with 10 and 100 ng or in case of 60-min incubation with 0.01, 0.1, 1, 10, and 100 ng. There was little difference in fluorescence between the negative control and all the other DNAs tested in 20- and 30-min incubations. On the other hand, the WBPH-65 primer set without LF and LB primers showed little amplification in the genomic DNAs of the three rice planthoppers, S. furcifera, N. lugens, and L. striatellus in a 60-min incubation. These results suggest that all six primers (F3, B3, FIP, BIP, LF, and BF) are necessary for the WBPH-65 primer set to detect S. furcifera within a 60-min incubation, and is able to discriminate S. furcifera from at least N. lugens and L. striatellus.

Development of an Efficient Method of Screening for Watermelon Plants Resistant to Fusarium oxysporum f. sp. niveum (수박 덩굴쪼김병에 대한 효율적인 저항성 검정법 개발)

  • Jo, Eun Ju;Lee, Ji Hyun;Choi, Yong Ho;Kim, Jin-Cheol;Choi, Gyung Ja
    • Horticultural Science & Technology
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    • v.33 no.3
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    • pp.409-419
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    • 2015
  • This study was conducted to establish an efficient screening method for watermelon plants resistant to Fusarium wilt (FW), which is caused by Fusarium oxysporum f. sp. niveum (Fon). An HA isolate was prepared from a wilted watermelon plant in Haman-gun and identified as F. oxysporum f. sp. niveum based on morphological characteristics, molecular analyses of ITS (internal transcribed spacer) and TEF (translation elongation factor $1{\alpha}$) sequences, and host specificity on cucurbits including watermelon, melon, oriental melon, and cucumber. The assay for disease response of watermelon differentials indicated that the HA isolate was race 0. Among seven liquid media tested, the highest amount of Fon spores was produced from V8-juice broth, which was selected as a medium for mass production of Fon. The disease assay for 21 watermelon and 11 watermelon-rootstock cultivars demonstrated that 20 watermelon cultivars except for 'Soknoranggul' were susceptible; 'Soknoranggul' was moderately resistant. All the tested rootstock cultivars were highly resistant to the HA isolate. The evaluation of disease development depending on various conditions suggested that an efficient screening method for FW resistance in watermelon plants is to dip the roots of 10-day-old seedlings in spore suspension of $1.0{\times}10^5-1.0{\times}10^6conidia{\cdot}mL^{-1}$ for 30 min., to transplant the seedlings to plastic pots with a fertilized soil, and then to cultivate the plants at $25^{\circ}C$ for 3 weeks.

Development of an Efficient Screening System for Resistance of Watermelon Plants to Didymella bryoniae (수박 덩굴마름병에 대한 효율적인 저항성 검정 방법 개발)

  • Lee, Ji Hyun;Jang, Kyoung Soo;Choi, Yong Ho;Kim, Jin-Cheol;Choi, Gyung Ja
    • Research in Plant Disease
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    • v.22 no.2
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    • pp.72-80
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    • 2016
  • Gummy stem blight, caused by the fungus Didymella bryoniae, is major disease of watermelons worldwide. The objective of the present study was to establish an efficient screening system to identify watermelon resistant to D. bryoniae. An GSB3 isolate was prepared from a watermelon plant showing typical symptoms of gummy stem blight in Haman-gun and identified as D. bryoniae based on molecular analysis of internal transcribed spacer sequence. A simple mass-production technique of inoculum was developed based on spore production of D. bryoniae GSB3 under several incubation conditions and their virulence on watermelon plants. Resistance degrees of 22 commercial watermelon cultivars to the GSB3 isolate were evaluated. Among them, four watermelon cultivars showing different degree of resistance response were selected for further study. Development of disease on the cultivars according to various conditions including inoculum concentrations, incubation periods in dew chamber, and incubation temperatures was investigated. From the results, we suggest an efficient screening method for resistant watermelon cultivars to gummy stem blight. Seeds of watermelon cultivar are sown and grown in a greenhouse until plant stage of 2-fully expanded leaves. Seedlings are inoculated with D. bryoniae by spraying spore suspension of the fungus at a concentration of $5.0{\times}10^5spores/ml$. The infected plants are incubated in humidity chamber at $25^{\circ}C$ for 48 hours and then transferred to a growth chamber at $25^{\circ}C$ and 80% relative humidity with 12-hour light a day. Three to four days after inoculation, disease severity of the plant are measured using percentage of infected leaf area.

Application of Multiplex RT-PCR for Simultaneous Identification of Tomato Spotted Wilt Virus and Thrips Species in an Individual Thrips on Chrysanthemum (시설재배 국화에서 총채벌레의 종 동정 및 보독 바이러스 동시 검출을 위한 다중 진단법 적용)

  • Yoon, Ju-Yeon;Yoon, Jung-Beom;Seo, Mi-Hye;Choi, Seung-Kook;Cho, In-Sook;Chung, Bong-Nam;Yang, Chang Yeol;Gangireddygari, Venkata Subba Reddy
    • Research in Plant Disease
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    • v.26 no.4
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    • pp.264-271
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    • 2020
  • We have developed a simultaneous diagnostic method that can identify both the species of thrips and tomato spotted wilt virus (TSWV) that are problematic in chrysanthemum plants. This is a method of amplifying DNA by performing reverse transcription-polymerase chain reaction by simultaneously adding primers specific to TSWV coat protein (N) gene and primers specific to the internal transcribed spacer 2 region of Frankliniella occidentalis and F. intonsa using total nucleic acid extracted from one thrips. The sizes of DNA fragments for TSWV, F. occidentalis, and F. intonsa were 777, 287, and 367 bp, respectively. These results showed species identification of thrips and whether thrips carrying TSWV can be simultaneously confirmed. Further usefulness of the simultaneous diagnostic method was made from greenhouse survey at chrysanthemum greenhouses in Taean (Chungcheongnam-do) and Changwon (Gyeongsangnam-do) to investigate the identification of thrips species and the rate of thrips carrying TSWV. Of thrips collected from the greenhouses, 83.7% thrips was F. occidentalis and 72.9% F. occidentalis carried TSWV in Taean. Similarly, the diagnostic method showed that 92.2% thrips was F. occidentalis and 84.0% F. occidentalis carried TSWV in Changwon. These results confirm that F. occidentalis is a dominant thrips species and the thrips species plays a crucial role in the transmission of TSWV in chrysanthemum plants in the greenhouses. Taken together, this study showed a simple diagnostic method for thrips identification and epidemiological studies of the timing and spread of TSWV through thrips in chrysanthemum greenhouses in South Korea.

Identification and Chemotype Profiling of Fusarium Head Blight Disease in Triticale (국내 재배 트리티케일에 발생한 붉은곰팡이병의 다양성 및 독소화학형 분석)

  • Yang, Jung-Wook;Kim, Joo-Yeon;Lee, Mi-Rang;Kang, In-Jeong;Jeong, Jung-Hyun;Park, Myoung Ryoul;Ku, Ja-Hwan;Kim, Wook-Han
    • Research in Plant Disease
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    • v.27 no.4
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    • pp.172-179
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    • 2021
  • This study aimed to assess the disease incidence and distribution of toxigenic in Korean triticale. The pathogen of triticale that cause Fusarium head blight were isolated from five different triticale cultivars that cultivated in Suwon Korea at 2021 year. The 72 candidate were classified as a Fusarium asiaticum by morphology analysis and by ITS1, TEF-1α gene sequence analysis. And the results of pathogenicity with 72 isolates on seedling triticale, 71 isolates were showed disease symptom. Also, seven out of 71 Fusarium isolates were inoculated on the wheat, to test the pathogenicity on the different host. The results showed more low pathogenicity on the wheat than triticale. The results of analysis of toxin type with 72 isolates, 64.6% isolates were produced nivalenol type toxin and other 4.6% and 30.8% isolates were produce 3-acetyldeoxynivalenol and 15-acetyldeoxynivalenol, respectively. To select fungicide for control, the 72 Fusarium isolates were cultivated on the media that containing four kinds fungicide. The captan, hexaconazole, and difenoconazole·propiconazole treated Fusarium isolates were not showed resistance response against each fungicide. However, six isolates out of 72 isolates, showed resistance response to fludioxonil. This study is first report that F. asiaticum causes Fusarium head blight disease of triticale in Korea.

Isolation and Identification of Competitive Fungi on Medium for Black Wood Ear Mushroom in Korea and In Vitro Selection of Potential Biocontrol Agents (목이버섯 배지 오염 곰팡이균의 분리, 동정 및 생물학적 방제제 선발)

  • Seoyeon Kim;Miju Jo;Sunmin An;Jiyoon Park;Jiwon Park;Sungkook Hong;Jiwoo Kim;Juhoon Cha;Yujin Roh;Da Som Kim;Mi jin Jeon;Won-Jae Chi;Sook-Young Park
    • Research in Plant Disease
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    • v.30 no.1
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    • pp.66-77
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    • 2024
  • Black wood ear mushroom (Auricularia auricula-judae) is one of the most economically important mushrooms in China, Japan, and Korea. The cultivation of wood ear mushrooms on artificial substrates is more efficient in terms of time and cost compared with their natural growth on trees. However, if the substrate cultivation is infected by fast-growing fungi, the relatively slow-growing ear mushroom will be outcompeted, leading to economic losses. In this study, we investigated the competitive fungal isolates from substrates infected with fast-growing fungi for the cultivation of ear mushrooms in Jangheung and Sunchon, Korea. We collected 54 isolates and identified them by sequencing their internal transcribed spacer region with morphological identification. Among the isolates, the dominant isolates were Trichoderma spp. (92.6%), Penicillium spp. (5.6%), and Talaromyces sp. (1.8%). To find an appropriate eco-friendly biocontrol agent, we used five Streptomyces spp. and Benomyl, as controls against Trichoderma spp. and Penicillium spp. Among the six Streptomyces spp., Streptomyces sp. JC203-3 effectively controlled the fungi Trichoderma spp. and Penicillium spp., which pose a significant problem for the substrates of black wood ear mushrooms. This result indicated that this Streptomyces sp. JC203-3 can be used as biocontrol agents to protect against Trichoderma and Penicillium spp.