|Goodman, Cynthia - Cindy|
Submitted to: BioProcessing
Publication Type: Peer reviewed journal
Publication Acceptance Date: 8/4/2005
Publication Date: 9/14/2005
Citation: Mcintosh, A.H., Grasela, J.J., Goodman, C.L. 2005. Simplified and rapid method for the extraction of DNA from baculovirus occlusion bodies. BioProcessing J. 5(2):59-61. Interpretive Summary: This study was initiated in order to develop a more efficient method for the extraction of DNA from specific insect viruses. These viruses, called baculoviruses, are used in the control of insect pests of agricultural importance. When they infect the insects, large numbers of spherical-like bodies containing the viral particles are produced. DNA is extracted from these viral particles in order to study their biological properties. The method described in this report was very efficient in recovering viral DNA from low numbers of the spherical-like bodies compared to other conventional methods. The impact of such a study will benefit both academic and industrial research, and will aid in the production of products of medical and agricultural importance.
Technical Abstract: The present report describes a simple convenient method for recovering DNA from occlusion bodies (OB) of baculoviruses. Compared with conventional methods, fewer OB (2x10*7) were required for the extraction of sufficient amounts of DNA (59ug) for molecular biology studies. The extracted DNA was capable of transfecting Trichoplusia ni (TN-CL1) cells with concomitant production of OB and extracellular virus (ECV). Restriction endonuclease analysis (REN) of the DNA extracted by the modified method gave identical profiles to the DNA extracted by conventional methods, thus establishing the integrity of the extracted DNA. Occlusion bodies can be produced in either cell culture, as was done in the present report, or in susceptible larvae. TN-CL1 cells were highly susceptible to AcMNPV (Ac-HPP) and produced 150 OB per cell. Once the OB were produced they were purified on sucrose gradients and DNA extracted by the method as described in this report. The entire process following purification of OB was carried out in 1.5 ml microcentrifuge tubes.