Identification of Citrus Exocortis Viroid by RT-PCR and Dot-Blot Hybridization

Year 2003, Volume: 16 Issue: 2, 221 - 228, 01.12.2003

M. Göçmen T. Taşdemir M. Kelten S. Güneş İ. Polat

Abstract

Citrus Exocortis Viroid (CEV) which has low weight and RNA structure 371 nucleotid lenght is very common in all citrus orchards. Its economic loss is much more on orange, mandarine, tangelo and grapefruid grafted especially on Poncirus trifoliata (L.), Carrizo citrange ve Rangpur lime rootstocks. Biological testing is the most common method for identification of CEV. This method takes long time and needs controlled greenhouse conditions, therefore the advanced molecular tests can be utilized to determine CEV. In this study, RT-PCR (Reverse Transkiriptase-Polymeraze Chain Reaction), a molecular technique identifying at RNA level and dot-blot hybridization via enzym (Biotin-ULS) marking technique was used for identification of CEV in Citrus Variety Improvement Programme in Turkey. In this way, presence of CEV in citrus has been identified more reliable and faster than biological testing method.

Keywords

Citrus, Exocortis Viroid, RT-PCR (Reverse Transkiriptase-Polymeraze Chain Reaction), Dot-Blotting hybridization

Turunçgil Exocortis Viroidinin RT-PCR ve Dot Blot Hibridizasyon Yöntemiyle Tanısı

Abstract

Turunçgil Exocortis viroidi (CEV), düşük moleküler ağırlıklı, 371 nükleotid uzunluğunda RNA yapısında olup, dünyada turunçgil yetiştiriciliği yapılan tüm alanlarda mevcuttur. Poncirus trifoliata (L.) Raf anacı (üç yapraklı) başta olmak üzere Carrizo citrange ve Rangpur lime üzerine aşılı portakal, mandarin, tangelo ve altıntopta ekonomik zararı fazladır. Turunçgil Exocortis hastalığının tanısında en yaygın kullanılan yöntem, biyolojik testlemedir. Ancak bu testleme yönteminde, kontrollü sera gerekliliği ve testleme süresinin uzun olması yeni modern tekniklerin kullanılmasını zorunlu kılmaktadır.Yapılan bu çalışmada, moleküler tekniklerden olan ve RNA düzeyinde tanılama yapmaya imkan sağlayan RT-PCR (Reverse Transkiriptase-Polymeraze Chain Reaction), ve enzim (Biotin-ULS) işaretlemesiyle Dot-Blot Hibridizasyon teknikleri Türkiye Turunçgil Çeşit Geliştirme Programında (TTÇGP) CEV’in tanılamasında kullanılmıştır. Böylece, biyolojik testleme süresi uzun olan CEV’nin, kısa sürede, güvenilir ve hızlı şekilde tanılamanın yapılabileceği görülmüştür.

Keywords

Turunçgil, Exocortis Viroid, RT-PCR (Reverse Transkiriptase-Polymeraze Chain Reaction), Dot-Blot Hibridizasyon

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