Nucleic Acids Research, 1990, Vol. 18, No. 19 5667-5672
© 1990
MOLECULAR BIOLOGY |
Linearization of baculovirus DNA enhances the recovery of recombinant virus expression vectors
NERC Institute of Virology and Environmental Microbiology Mansfield Road, Oxford OX1 3SR, UK
*To whom correspondence should be addressed
Received July 26, 1990. Revised September 3, 1990. Accepted September 3, 1990.
Engineered derivatives of Autographs californica multiple nucleocapsid nuclear polyhedrosis virus (AcMNPV) possessing a unique restriction site provide a source of viral DNA that can be linearized by digestion with a specific endonuclease. Circular or linearized DNA from two such viruses were compared in terms of their infectivity and recombinogenic activities. The linear forms were 15- to 150-fold less infectious than the corresponding circular forms, when transfected into Spodoptera fruglperda cells using the calcium phosphate method. Linear viral DNA was, however, proficient at recombination on co-transfection with an appropriate transfer vector. Up to 30% of the progeny viruses were recombinant, a 10-fold higher fraction of recombinants than was obtained from co-transfections with circular AcMNPV DNA. The isolation of a recombinant baculovirus expression vector from any of the AcMNPV transfer vectors currently In use can thus be facilitated by linearization of the viral DNA at the appropriate location.
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