Korean Journal of Plant Tissue Culture (식물조직배양학회지)

The Korean Society of Plant Biotechnology (한국식물생명공학회)
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Improved Detection and Purification of Grapevine Leafroll-associated 3 Closterovirus Using Tissue Culture

포도 조직배양에 의한 Grapevine Leafroll-associated 3 Closterovirus의 증식과 검출효율 증대

  • Published : 2001.11.01
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Abstract

Grapevine leafroll-associated 3 closterovirus (GLRaV-3) is phloem limited virus, and one of the most severe viral diseases found in Korea. However, nonhomogenous distribution and low concentration and seasonal variations of GLRaV-3 in grapevines remain as main problems which prevent the introduction and molecular biology or serology experiments. Virus-infected plantlet in vitro was obtained from node tissue cuttings, which was GLRaV-3 infected 'kyoho' vines. The amount of purified virus was highest in vitro plantlet. Moreover, viruses seem to be relatively homogeneously distributed in all organs including leaf, stem and callus derived from in vitro plantlets. RT-PCR detection using in vitro plantlet tissue as template was most effective. When comparing ELISA to RT-PCR, RT-PCR detection was 1,000 times as effective as ELISA. These results can be explained by improved quality such as tenderness or less tannins in plantlet in vitro. In conclusion, until infected herbaceous host will be available, tissue culture can be usefully adopted as a technique for a good source of GLRaV-3 closterovirus for further studies.

우리나라 포도원에서 가장 많이 발생하여 피해를 주고 있는 grapevine leafroll-associated 3 Closterovirus (GLRaV-3)를 대상으로 기내 배양묘를 이용한 바이러스 순화 및 진단효율성을 검토하였다. 바이러스 감염주의 절간을 기내배양하여 증식시킨 기내 배양묘를 1개월 간격으로 계대배양하면서 배양묘를 ELISA 검정한 결과 고농도의 바이러스가 검출되었으며 배양묘의 부위별로 잎, 줄기 및 줄기 유래의 callus 조직에서 모두 고농도로 검출되었다. 또한 포장에서 재배되고 있는 포도나무의 잎이나 엽병조직에 비해 기내 배양묘의 조직을 사용하였을 때 높은 농도의 정제 바이러스를 얻을 수 있었으며, 전자현미경에 의한 dip 검경에서도 사상형 바이러스 입자가 관찰되었다. RT-PCR 진단에서는 기내 배양묘 조직을 사용하였을 때 포도 유엽조직과 엽병+중륵조직에 비해 검출효율이 높았다. 기내 배양묘의 잎조직을 이용하여 ELISA와 RT-PCR 검정감도를 비교한 결과 ELISA에 비해 RT-PCR 검정이 약 1,000배 감도가 높았다.

Keywords

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