Design of a low-cost process for peroxidase production in insect larvae

LUCÍA V. ROMERO; ALEXANDRA M. TARGOVNIK; GUSTAVO J. LEVIN; MARÍA DE LAS NIEVES LOUSTAU; OSCAR TABOGA; OSVALDO CASCONE; MARIA V. MIRANDA

Mar del Plata, Buenos Aires. Argentina

Congreso; XLIII Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2007

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular.

Baculovirus-insect cell system is a well-known choice for expressing foreign genes. We describe a technology to scale up horseradish peroxidase isozyme C (HRPC) production using insect larvae as biofactories. Two strategies were applied in order to obtain polyhedra, the natural viral infective structure, containing rAcMNPV: 1-Mixed polyhedra (MP) were obtained by co-infection of wild type AcMNPV and AcMNPV HRP+/occ- at different multiplicities of infection (MOI) of both virus and ratios between them. 2- AcMNPV HRP+/occ+ construction by co-transfection with transfer vector pAcGP67-HRP and bAcGOZA. Four instar starved larvae of R. nu and S. frugiperda were fed with artificial diet contaminated with 1x106 pol/100 mg of both polyhedra suspensions and HRP activity was measured in haemolymph at days 4 and 8 post-infection respectively.R. nu larvae infected with MP containing 98.9% rAcMNPV (as determined by real time PCR) expressed 31.1 mg HRPC/l. When the same larvae were infected with AcMNPV HRP+/occ+ polyhedra, HRPC production was approximately 50% higher (46.8 mg/l). Similar results were obtained with S. frugiperda larvae but HRPC production was 137 lower. Taking these results into account, R. nu larva is the best choice for HRPC production by oral infection with AcMNPV HRP+/occ+. This low-cost strategy is a practical way for infecting a high number of insects easily