Insect cell extract preparation by the nitrogen disruption method for cell-free translation

An insect cell extract for cell-free translation was prepared using a Mini- Bomb cell disruption chamber, a unique device which disrupts biological mat erials by altering the pressure on these materials. The highest potential e xtract for cell-free translation was prepared from the cell suspension at a density of 1.0 X 10(8) cells/ml, then disrupted at a nitrogen pressure of 10 kg/cm(2). Furthermore, synthesized proteins were stable in this extract, but were degraded by proteinase in an extract prepared with a Potter-Elveh jem homogenizer. The insect cell extract showed efficient activity for tran slation of protein 2a and coat protein of the brome mosaic virus, while ine fficiently translating protein la and protein 3a. On the other hand, mainly protein 3a and coat protein were translated in a wheat germ system, while mainly proteins 2a and 3a were translated in a rabbit reticulocyte system. The insect cell-free system appears to possess an unusual property for enha ncing translation of exogenous mRNA, making it different from the rabbit re ticulocyte and wheat germ systems.