produção de sintomas e movimento seguido de agro-inoculação com ssdna de “tomato yellow leaf...

Produção de sintomas e movimentoseguido de agro-inoculação com

ssDNA de “tomato yellow leaf curlvirus” Geminivirus (Tailândia).

Instituto Federal Goiano câmpus Urutaí.Curso de Agronomia

Disciplina de Fitopatologia I

Apresentador: Tony L. Moura

ROCHESTER, D.E., KOSITRATANA, W., BEACHY., R.N. Systemic

Movement and Symptom Production following Agroinoculation with a

Single DNA of Tomato Yellow Leaf Curl Geminivirus (Thailand).

VIROLOGY 178,520-526 (1990) 1

INTRODUÇÃO• Vários gêneros pertencentes a família Geminiviridae

• Geminiviroses: Solanaceae, leguminosas e gramíneas;

• A espécie - TYLCV em diferentes regiões;

• Transmissão: cigarrinha e mosca branca;

• Suas sequências usadas para construção de árvoresfilogenéticas;

• Patógeno de grande importância para o tomate noMediterrâneo.

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OBJETIVO• Este estudo procura inicialmente caracterizar o

genoma de um TYLCV isolado da Tailândia,encontrado infectando tecido de tomate.

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MATERIAIS E MÉTODOS• A purificação do Vírus;

• Clonagem do DNA do genoma do TYLCV;

• Construção do clone por Agroinoculação;

• Agro-inoculação e análise de plantasinfectadas;

• Extração de DNA e análise;

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RESULTADOS E DISCUSSÃO

FIG. 1. As sequências das regiões comuns TYLCV: setas sólidas indicam repetiçõescomuns, setas tracejadas indicam a posição estável no caule e uma estrutura dehaste e laço estável pode formar. O oligonucleotideo (DR2) usado como um primesecundário abaixo da vertente síntese primordial abaixo da linha. Apenas diferençasde sequência de TYLCV-A TYLCV-B são mostrados. Asteriscos indicam posições ondeas lacunas são inseridas para proporcionar o máximo de homologia.

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FIG. 2. Southern blot de TYLCV recombinante (Tailândia) clones, pDR9 (putativo componenteA) e pDR7 (putativo componente B). 1, pDR7 (1 pg), digerido com EcoRI; 2. pDR7 (1 pg),digerido com EcoRI e Sphl; 3. pDR9 (0,5 pg) digerido com EcoRI; 4. pDR9 (1 pg), digerido comEcoRI; 5. pDR9 (1 pg), digerido com EcoRI e BarnHI. 7


FIG. 3. TYLCV - Nicotiana benthamiana infectadas 3 semanas após agroinoculação. (a) Asplantas agro-inoculadas com “TYLCV-A" só, “TYLCV-B" sozinho, e “TYLCV A e B. t" (b) Plantaque foi agro-inoculada com apenas com o componente A (pDR21: SE). (c) Plantas inoculadascom "A" e "B" componentes (pDR2 1: SE e pDR37: SE, respectivamente). 8


Fig. 4. Análise de Dot blot de TYLCV encontrado infectando plantas de tomate. Tratamentos 1-4 - somente o componente “A”, 5-8 componente B, 9-12 componentes A e B, 13-16 a bactéria responsável pela agro-inoculação (A. tumefaciens 3111 SE), 17-20 não agro-inoculadas com TYLCV.

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RESULTADOS E DISCUSSÃO2

FIG. 5. Análise de Southern blot de plantas de Nicotiana benthamiana infectada por TYLCV. Na (A) linha 1, inoculada com A. tumefaciens 3111SE; linha 2, , inoculada com A. tumefaciens do TYLCV-“B”; linha 3, agro-inoculado com TYLCV-“A”; linha 4, agro-inoculado com TYLCV- “A” (enzima “bean nuclease “) ; linhas 5 e 7, agro-inoculado com TYLCV-“A + B”; linhas 6 e 8, agro-inoculado com TYLCV-“A + B” (enzima “bean nuclease “). Linhas l-6 foram hibridizadas com sondas específicas de TYLCV-“A”-e linhas 7, 8, com sondas específicas de TYLCV-“B”. Tipos de DNA : OC, open circle; L, linear; CCC, covalently closed circle; SS, single-stranded.

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CONCLUSÕES

• Verificou-se que o TYLCV - é um DNA capaz de infectar através de agroinoculação, mover sistemicamente, e induzir sintomas como mal-estar, tanto tomate e N. benthamiana.

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produção de sintomas e movimento seguido de agro-inoculação com ssdna de “tomato yellow leaf...

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