Limpeza clonal de variedades de mandioca (Manihot esculenta Crantz) e produção de antissoro para o vírus do mosaico comum da mandioca

• Autor(a) principal: Carnelossi, Patrícia Rosin
• Data de Publicação: 2010
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Estadual de Maringá (RI-UEM)
• Texto Completo: http://repositorio.uem.br:8080/jspui/handle/1/1265
• topic: Mandioca (Manihot esculenta Crantz); Vírus do mosaico comum (CsCMV); Cultura de meristemas; Termoterapia; Teste de ELISA-indireto; Purificação; Produção de antissoro.; CsCMV; Meristem culture; Thermotherapy; Serology; Indirect-ELISA tests.; Ciências Agrárias; Agronomia

Part of the production of cassava (Manihot esculenta Crantz) is directed towards human feeding through consumption of its roots. Due to its vegetative propagation, it is subject to the dissemination of systemic diseases as those caused by viruses after successive generations. CsCMV is the most disseminated in the cassava producing areas of the world. In this work, a clonal cleaning protocol was evaluated in order to obtain cleaned plants free from CsCMV of the varieties of cassava (Olho Junto, Fécula Branca, IAC 90, IAC 12, IAC 13, IAC 14 e Baianinha) and the material "Pasquini" (non-defined variety) cultivated in the Northwest region of Parana State. A specific antiserum for CsCMV was produced and Indirect ELISA tests revealed the natural infection of CsCMV in all the cassava varieties tested. The micropropagation protocol was efficient for Olho Junto and IAC-12 varieties and also for "Pasquini", resulting in healthy plantlets, which were ready for acclimatization. For the other varieties it is necessary to adapt this method. The results showed that 31 out of 64 cassava plantlets developed in vitro proceeding from thermotherapy associated with meristem tip culture (independently from the variety) were free from CsCMV after indexation tests through ELISA. The addition of Ribavirin to the MS culture medium at the concentrations of 5 - 40 mg/L was not efficient for the conditions tested, causing fitotoxic effects and death of the 3 varieties tested. The CsCMV purification was efficient and a highly specific antiserum with a high titre (1:10.000) was raised. When used in ELISA tests, the prepared purifications of specific G immunoglobulins (IgG) for CsCMV were equally successful and showed to be more sensitive to virus detection than the pure antiserum. The molecular mass of the purified CsCMV coat protein was determined, which was close to 20 KDa and within the expected value for Potexvirus. RT-PCR reactions using specific primers for Potexvirus resulted in the amplification of fragments (around 760 bp), which possibly represent the partial amplification of the viral replicase coding gene.