담배가루이(Bemisia tabaci, Aleyrodidae, Hemiptera)에서 Virus-induced Gene Silencing (VIGS) Vector를 이용하기 위한 cDNA Library 제작

고나연,임현섭,유용만,윤영남,Ko, Na Yeon,Lim, Hyoun Sub,Yu, Yong Man,Youn, Young Nam 한국응용곤충학회 2015 한국응용곤충학회지 Vol.54 No.2

담배가루이(Bemisia tabaci)는 외래해충으로 바이러스벡터로 작용하여, 토마토의 토마토황하잎말림병바이러스(TYLCV)를 비롯한 약 100여종의 바이러스를 매개하는 중요한 해충이다. 본 연구에서는 VIGS vector를 이용하여 담배가루이 방제를 위한 target 유전자들을 선발하기 위해 gateway system을 이용한 담배가루이 cDNA library 제작을 시도하였다. 첫 번째 방법으로 oligo d(T) primer를 사용하였을 때, 평균 약 1 kb의 insert와 $1.4{\times}10^4cfu$의 titer를 확인하였다. 그러나 insert size가 너무 커서 적절하지 않았다. 두 번째 방법으로 attB-N25 random primer를 이용하고, sonication을 6초 실시하여 다시 진행하였다. 그러나 확인되는 insert size는 다소 컸고, 몇몇은 insert가 너무 작아서 밴드가 확인 되지않았으며, $1.04{\times}10^54cfu$의 titer를 확인할 수 있었다. 세 번째 방법으로는 oligo d(T) primer를 이용하였고, sonication을 2초 실시하였다. 그 결과 300 bp~600 bp size의 insert가 확인되었으나, electro transformation을 사용한 첫번째, 두번째 방법에 비해 heat shock transformation을 사용하여 titer가 $5.2{\times}10^24cfu$로 매우 낮은 것을 확인 할 수 있었다. 결과적으로 cDNA library를 만들 때 먼저 random primer를 사용하여 First strand를 합성하여 poly A를 제거하고, 다음으로 sonication을 1초 실시하여 300~700 bp정도의 적절한 size의 insert를 생성하고, 마지막으로 electro-transformation을 실시하여 transformation 효율을 높인다면 VIGS vector에 적합한 cDNA library를 만들 수 있을 것으로 사료 된다. The sweetpotato whitefly, Bemisia tabaci, is the major insect pest that transmitted over 100 plant viruses including tomato yellow leaf curl virus (TYLCV) of tomato plant as virus vector in the world. In this study, cDNA library of whitefly was constructed using Gateway system for selecting target gene in order to control of B. tabaci using virus-induced gene silencing (VIGS) vector with RNAi. First of all, when using oligo d(T) rimer, the calculated titer of cDNA library was confirmed with $1.4{\times}10^4$ clones and average insert sizes was confirmed with 1 kb. However, insert size was very big for construction of cDNA. Otherwise, when using attB-N25 random primer and sonication for 6 sec, the calculated titer of cDNA library was confirmed with $1.04{\times}10^5$ clones. But mostly insert band wasn't identified on the electrophoresis, because it seemed that insert size is too small (${\leq}100bp$), also the size of identified insert was somewhat big. Finally, when using oligo d(T) primer and sonication for 1 sec, cDNA insert of whitefly was appropriated for VIGS with 300-600 bp. However, cDNA sequence included a poly A and titer was very low to $5.2{\times}10^2$ clones. It was supposed that heat shock transformation was used instead of electro-transformation. It is considered that when constructing cDNA library for using VIGS vector, (1) random primer should be used for First strand cDNA synthesis in order to remove poly A and (2) sonication for 1 sec should be performed in order to get appropriated insert size and (3) electro-transformation should be performed in order to improve transformation efficiency.

RNAi에 의한 담배가루이(Bemisia tabaci, 가루이과, 노린재목)의 개체군 밀도변화

고나연(Na-Yeon Ko),윤영남(Young-Nam Youn) 충남대학교 농업과학연구소 2015 농업과학연구 Vol.42 No.1

Ninety genes randomly selected from tobacco whitefly (Bemisia tabaci) cDNA library was studied for selecting target gene in order to control of tobacco whitefly using TRV-VIGS vector (tobacco rattle virus-virus induced gene silencing vector) with RNAi. First of all, the occurrence of B. tabaci adult according to agro-infiltration of TRV was no significant difference. And that of TRV inserted tobacco whitefly cDNA showed a significant difference in each sample. P CV and N CV sample were more than 80% could be confirmed in 5 samples, for example, wh11, wh36, wh46, wh50 and wh71. Lastly, the occurrence of nymph and egg also showed a significant difference in each sample. That could be confirmed in 11 samples, for example, wh01, wh09, wh10, wh15, wh16, wh23, wh24, wh48, wh64 and wh66. In case of wh46, wh50 and wh71 sample could be confirmed that occurrence of B. tabaci adult was many, but occurrence of B. tabaci nymph and egg was a little. So sample showed a physioecological good effect to control of whitefly need to be investigated variation of gene expression in whitefly body using qRT-PCR through individual test.

지역별 시설재배지에서 식물기생선충의 살선충제에 대한 밀도억제 효과 비교

김세희,박상은,고나연,류태희,신효섭,권혜리,서미자,유용만,윤영남,Kim, Sae-Hee,Park, Sang-Eun,Ko, Na-Yeon,Ryu, Tae-Hee,Shin, Heo-Seob,Kwon, Hye-Ri,Seo, Mi-Ja,Yu, Yong-Man,Youn, Young-Nam 한국응용곤충학회 2013 한국응용곤충학회지 Vol.52 No.3

과채류 재배지에서 발생하는 식물기생성 선충의 방제를 위해 충남 공주의 오이 재배지와 충남 부여와 논산, 경남 진주의 딸기 재배지, 전남곡성의 메론 재배지역에서 시설재배와 노지 재배를 하고 있는 농가토양을 채취하여 선충을 분리, 동정한 후 각 지역별 토양에서 abamectin 1.68% SC, cadusafos 3% GR, dazomet 98% GR, fosthiazate 30% SL, BA12011 SL 등 약제들의 선충에 대한 밀도억제 효과를 알아보기 위하여 실내실험과 포장실험을 실시하였다. 대부분의 지역에서 채취한 토양 시료에 분포하고 있는 식물기생성 선충의 평균밀도가 약제처리 후 감소하는 것으로 확인되었지만, 부여의 경우 fosthiazate 처리시 Pratylenchus spp.가, 곡성의 경우 fosthiazate, abamectin, cadusafos, dazomet 처리시 Pratylenchus spp.에 대하여 토양 내 평균 밀도가 상승하였다. 또한 BA12011 처리시 약제처리 후 다른 약제와 비슷한 수준의 개체군 평균밀도를 보였으며, 특히 공주 토양에서는 fosthiazate의 약제처리 효과보다 모든 종에서 높은 방제효과를 확인하였다. 각 지역에서 채취한 토양에 약제처리를 한 후 난낭 형성을 확인한 결과, abamectin의 경우 부여와 진주의 토양에서 효과를 보였고, BA12011은 논산과 곡성, dazomet는 공주에서 난낭 형성을 억제하는 효과를 확인하였다. BA12011을 가지고 오이재배지에서의 방제효과를 살펴본 결과, Meloidogyne spp., Pratylenchus spp., Helicotylenchus spp.의 평균 밀도가 감소하였고, 2차 처리시보다 1차 처리시 평균 밀도의 감소율이 높은 것으로 보아 초기 단계의 방제의 중요성을 확인할 수 있었다. To assess the efficacy of nematicides for the control of plant-parasitic nematodes in fruit and vegetables fields, soil samples were collected from a cucumber field at Gongju; from strawberry fields at Buyeo, Nonsan, and Jinju; and from a melon field at Gocksung in Jeonnam Province, Korea. Plant-parasitic nematodes were separated from each soil sample and identified. The susceptibilities of the nematodes to abamectin 1.68% SC, cadusafos 3% GR, dazomet 98% GR, fosthiazate 30% SL and BA12011 SL were examined under laboratory and field conditions. The average population density of plant-parasitic nematodes was generally reduced after the treatment with nematicides; however, there was increase in the population of Pratylenchus spp. in soil after treatment with fosthiazate at Buyeo and Gocksung. Furthermore, there were increased populations following treatment with abamectin, cadusafos, and dazomet at Gocksung. The control effects of BA12011 treatment on plant-parasitic nematodes were confirmed to be similar to those of the other 4 nematicides evaluated, although its control effect was higher than that of fosthiazate in cucumber-growing soil at Gongju. The effects of nematicide treatment on egg mass formation in each of the collected soils differed according to the region of soil origin. Abamectin was effective in reducing the degree of egg mass formation in Buyeo and Jinju soil, whereas BA12011 was effective in Nonsan and Gocksung soil. Dazomet was found to inhibit egg mass formation in Gongju soil. To evaluate the effect of the newly developed nematicide, BA12011, experiments were conducted in a cucumber-growing greenhouse. The average population densities of Meloidogyne spp., Pratylenchus spp., and Helicotylenchus spp. after the first treatment were reduced to a greater extent than after the second treatment. It is thus suggested that early nematicide treatment is important for effective control of plant-parasitic nematodes.

하우스시설 재배지에서 발생하는 주요 선충

김세희(Sae-Hee Kim),박상은(Sang-Eun Park),고나연(Na-Yeon Ko),류태희(Tae-Hee Ryu),신효섭(Heo-Seob Shin),권혜리(Hye-Ri Kwon),서미자(Mi-Ja Seo),유용만(Yong-Man Yu),윤영남(Young-Nam Youn) 충남대학교 농업과학연구소 2013 농업과학연구 Vol.40 No.2

To know how much damages occurred by nematodes in plastic vinyl house field, soil samples were collected from strawberry cultivation areas at Buyeo and Nonsan in Chungnam and Jinju in Kyeongnam, melon cultivation area at Gocksung in Junnam and cucumber field at Gongju in Chungnam. And then, nematode samples were separated from each soil sample, and identified the kind of plant-parasitic nematodes. Plant-parasitic nematodes were separated from 52 soil samples. Among samples, Meloidogyne spp., Pratylenchus spp. and Helicotylenchus spp. were isolated from 45 (86.5%%) and 33 (63.5%) and 47 collected soil samples (90.4%), respectively. As a result of identification of plant-parasitic nematodes from regional collected soil samples, distribution of Helicotylenchus spp. was higher than any other plant-parasitic nematode. And the population of Meloidogyne spp. and Pratylenchu spp. were also higher, and a occurrence ratio of Meloidogyne spp. is higher than Pratylenchu spp. except the cucumber growing area at Gongju.

구기자에서 큰이십팔점박이무당벌레(Henosepilachna vigintioctomaculata)의 온도조건에 따른 발육특성과 친환경제재에 의한 방제효과

강찬영(Chan-Yeong Kang),류태희(Tae-Hee Ryu),정유빈(Yu-Bin Jung),고나연(Na-Yeon Ko),권혜리(Hye-Ri Kwon),서미자(Mi-Ja Seo),유용만(Yong-Man Yu),윤영남(Young-Nam Youn),김영국(Yeong-Guk Kim) 충남대학교 농업과학연구소 2014 농업과학연구 Vol.41 No.4

Host plants of the Henosepilachna vigintioctomaculata are known as Solanaceae plants such as potato, tomato, eggplant and Lycium chinense. Henosepilachna vigintioctomaculata (Motschulsky) cause continuous damage from early June to mid-August in leaves of L. chinense under environmental-friendly management in Cheongyang, Chungnam. The developmental period of egg and larva of H. vigintioctomaculata were investigated. H. vigintioctomaculata were reared on L. chinense at several temperatures, for example, 15, 20, 25 and 30°C in the laboratory. In addition, the control effect of 5 eco-friendly agricultural materials were examined. As results, hatching rate is the highest to 91% at 25°C. Egg periods were 10.8, 7.7, 5.1 and 3.7 days at each tested temperature, respectively. And the total developmental periods from egg to adult were 42.7, 26.3, 18.4 and 19.4 days at each tested temperature, respectively. Otherwise, their eclosion rates from pupa to adult were showed 58%, 77%, 85%, 65% at each tested temperature, respectively. As a result of the control effect, both adults and larvae of H. vigintioctomaculata were shown a high mortality to mixture with extract of Sophora flavescens, Chenopodium ambrosioides and Melia azedarach. Additional, larvae is shown high mortality to extract of Azadirachta indica and Cymbopogon nardus. Thus H. vigintioctomaculata might be controled by using plant extract of eco-friendly agricultural materials in due season.

친환경 구기자재배지에서 다년간 해충발생양상

류태희(Tae-Hee Ryu),강찬영(Chan-Yeong Kang),정유빈(Yu-Bin Jung),고나연(Na-Yeon Ko),권혜리(Hye-Ri Kwon),서미자(Mi-Ja Seo),유용만(Yong-Man Yu),윤영남(Young-Nam Youn),김영국(Yeong-Guk Kim) 충남대학교 농업과학연구소 2014 농업과학연구 Vol.41 No.4

The occurrence of insect pests of Lycium chinense was monitored at 4 organic farms that was cultivated with environment-friendly management at Cheongyang, Chungnam from May, 2012 to September, 2014. Total 4 orders and 18 species occurred in the L. chinense. Among them, 4 major pests and Lepidopteran and Hemipteran insect pests were investigated the occurrence pattern. During the budding, the population density of Myzus persicae was very high in late-May and early-June, and the injury by Lema decempunctata was increased from mid-June. The injuries by Henosepilachna vigintioctomaculata and Hedma lycia sp. were continued throughout the growing season. From the fruit held in late-July, hemipteran insects including Plautia stali and Halyomorpba balys were sucking the fruit, and their injury to L. chinense were gradually increased. In the case of Eriophys macrodonis densities to increase gradually from early July. The population density of E. macrodonis in 2014 was rapidly increased more than 5 times compared with 2012 and 2013. It was supposed that the initial control of mites or the removing of over-wintering density in the past year might be neglected. However, there is a need to more review the factors for the mass occurs.

친환경 구기자재배지에서 해충의 계절 발생소장 및 친환경유기농자재의 방제효과

류태희(Tae-Hee Ryu),박상은(Sang-Eun Park),고나연(Na-Yeon Ko),김정곤(Jeong-Gon Kim),신효섭(Heo-Seob Shin),권혜리(Hye-Ri Kwon),김영국(Yeong-Guk Kim),이보희(Bo-Hee Lee),서미자(Mi-Ja Seo),유용만(Yong-Man Yu),윤영남(Young-Nam Youn) 한국농약과학회 2013 농약과학회지 Vol.17 No.4