Submitted to: International Organization of Citrus Virologists Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 11/7/2004
Publication Date: 11/30/2004
Citation: Rangel, E., Manjunath, K.L., Guerra-Moreno, A.S., Brlansky, R.H., Lee, R.F. 2004. Bacterial expression and purification of a protein from citrus leprosis virus. International Organization of Citrus Virologists Abstracts, Page 95. Interpretive Summary:
Technical Abstract: Technical Abstract: Citrus leprosis is a serious disease in several countries in South America and the Caribbean basin. The disease is transmitted by Brevipalpus mites. At present, the identification of the disease is based mainly on visual symptoms followed by confirmation of presence of virus particles using transmission electron microscopy. Early identification of the disease from plant materials and mites is important for management of the disease and to minimize spread into new areas. A highly expressed protein from the putative cytoplasmic leprosis virus genome was cloned in a bacterial expression vector, pET 27b (Novagen). The leprosis protein with a carboxy-terminal fusion of a HSV and 6x Histidine tags was expressed in the expression host, Escherichia coli, strain BL21. The expressed protein was purified by two methods; the insoluble inclusion protein was purified by using Bugbuster (Novagen), and the soluble fraction protein was purified using an Ni-NTA agarose column. The expressed protein was monitored through different steps of purification by Western blotting using an anti-HSV monoclonal antibody. Both the expressed protein preparations were used as inject antigens in rabbits and chickens for producing antibodies to CiLV.