Analysis of proteins associated with storage root formation in cassava using two-dimensional gel electrophoresis

• Published in: Revista brasileira de fisiologia vegetal 2001, Vol.13 (1), p.41-48
• Main Authors: CABRAL, GLAUCIA B.(EMBRAPA-Genetic Resources and Biotechnology), CARVALHO, LUIZ J.C.B.(EMBRAPA-Genetic Resources and Biotechnology)
• Format: Article
• Language: English
• Subjects: 2-D gel electrophoresis; 2-DE géis; cassava; mandioca; Manihot esculenta; PLANT SCIENCES; proteins; proteínas; raízes de reserva; storage root
• ISSN: 0103-3131; 0103-3131
• DOI: 10.1590/S0103-31312001000100005

Two-dimensional gel electrophoresis analysis was performed on adventitious and storage root in cassava (Manihot esculenta Crantz). Adventitious root lacking swelling formation and swelled storage root were obtained from the accession WU104 grown in the green house of the Department of Biology in Washington University in St. Louis (MO-USA). Saline buffer-soluble proteins were extracted, separated in a high-resolution 2-D electrophoresis system, visualized with silver staining gel procedure, and digital image generate for further analysis. Quantitative and qualitative protein spots analysis was performed with a computer assisted image software system. Results revealed large variation in the complexity of the gel protein profile between the two root systems. About 90% of the protein spots appeared in the pI range value of 4.0 to 6.5 and between 14 to 80 Kda of molecular mass. Detailed computer assisted analysis of this gel allowed us to establish 5 distinct classes of protein based on spot quantification that could be associated with swelling and non-swelling roots. Variation in the complexity of protein pattern was related with different type of root. Whereas the adventitious root showed a more simple profile related to primary growth, the storage root showed to be a more complex profile related to secondary growth and starch accumulation. 2-DE géis foram usados para separar proteínas de raízes de reserva de mandioca (Manihot esculenta Crantz). Raízes adventícias desprovidas de engrossamento e raízes adventícias de reservas foram obtidas com o crescimento da cultivar WU104 em vasos em condições de casa-de-vegetação durante o verão de 1995 no Departamento de Biologia da Universidade de Washington em St. Louis (MO, USA). Proteínas solúveis em tampão fosfatado salino foram extraídas, separadas em um sistema de 2-DE de alta resolução, visualizadas com coloração em prata e utilizadas para gerar imagens digitais para análises computacionais posteriores. Análises quantitativos e qualitativos das proteínas foram proces-sadas em programas de computadores específicos. Os resultados revelaram uma grande variabilidade e complexidade nos perfis protéicos dos dois sistemas de raízes. Cerca de 90% das proteínas estavam distribuídas entre os valores de pI entre 4.0 a 6.5 e de peso molecular entre 14 a 80Kda de massa, formando padrões específicos para cada tipo de raiz. Com as analises computacionais detalhadas desses perfis protéicos diferenciados, foi possível estabelecer