[Purificacion y analisis de la proteina arcelina 5].

Food legumes constitute an important source of dietary protein for million of people throughout the world. In developing countries specially, cultivation of legumes is the best and quickest way to augment the production of food proteins. Limitation on availability of the legumes as food is due the tremendous losses caused by pests prior to and during storage. These pests include bruchid insect which attack different grain legumes. The mexican bean weevil, Zabrotes subfasciatus, and the bean weevil, Acanthoscelides obtectus, are the most important pests of stored beans (Phaseolus vulgaris). Cultivated Phaseolus vulgaris beans are infested by both species, although the prevalence or infestation by each species differs in different climatic zones. Damage of bruchids on beans is of considerable importance in Africa and throughout Latin America. It has been estimated that up to 35 percent of the crop in Mexico, Central America and Panama and 20-30 percent in Brazil is lost to bruchids infestations. Chemical control measure is used in attempting to minimize bruchid damage, but addition of a foreign material to a product near to consumption stages is often undesirable. Several wild forms of Phaseolus vulgaris of Mexican origin were reported presenting high levels of antibiosis resistance led to the hypothesis that the aracelin protein present in 10 percent of the wild Phaseolus vulgaris lines plays a role in the resistance to Z. subfasciatus. However, no information has been obtained for A. obtectus. Our main objective is to obtain cultivated dry beans resistant to two species of bruchids Z. subfasciatus and A. obtectus via genetic engineering and to express differents arcelin in microorganisms to be used in biological control of bruchids on stored beans. We have produced polyclonal and monoclonal antibodies against arcelin-1 and arcelin-5 a detailed study of immunocytolocalization was carried out. This study shows the presence of arcelin in the vacuolar protein body concentrated in cotyledon cells of wild bean seeds. The Arc1, Arc5 and Arc4 proteins were purified by using differential solubility properties and DEAE-trysacril ion exchange chromatography in order to use on antibiosis tests with both bruchids. We have constructed genomic libraries of different lines of wild Phaseolus vulgaris in order to isolate Arcelin genes using synthetic oligonucleotide as a probe. In addition, by using PCR technique the Arc1 and cDNA was isolated.