Infection, transfection, and co-transfection of baculoviruses by microprojectile bombardment of larvae.
The use of baculoviruses as expression vectors for heterologous proteins has been practically limited to the use of the Autographa californica multiple nucleopolyhedrovirus (AcMNPV). In this work, infection, transfection and co-transfection events with the baculoviruses AcMNPV and Trichoplusia ni granulovirus (TnGV) were accomplished by bombardment of T. ni first-instar larvae with microprojectiles coated with virions, viral DNA, and viral DNA and a transfer vector, respectively. A series of shooting conditions were tested until positive results were obtained. The use of 1.6 microm gold particles at 900 psi shooting pressure, 400 Torr vacuum, 7 cm distance to target, on sets of 20 first-instar larvae held in a 16 mm diameter container, proved to be the best shooting conditions. Typical infection symptoms were shown by larvae when shot with viruses or viral DNA from AcMNPV or TnGV. Co-transfected recombinant AcMNPV and TnGV were identified by the formation of occlusion bodies and GFP, respectively, in bombarded larvae. This technique opens a wide range of possibilities, not only to use an extensive number of baculoviruses as expression vectors for heterologous proteins, but also be used to infect, transfect or co-transfect a wide variety of viruses into animal cells.
Obregón-Barboza
Verónica
V
Departamento de Biotecnología y Bioquímica, Cinvestav, Irapuato, Gto., Mexico.
Del Rincón-Castro
Ma Cristina
MC
Cabrera-Ponce
José L
JL
Ibarra
Jorge E
JE
D000818:Animals; D001424:Bacterial Infections / Q000235:genetics / Q000821:virology*; D019470:Biolistics; D004279:DNA, Viral / Q000235:genetics; D005822:Genetic Vectors; D016679:Genome, Viral; D025722:Granulovirus / Q000235:genetics / Q000472:pathogenicity*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000821:virology; D007915:Lepidoptera / Q000235:genetics / Q000254:growth & development / Q000821:virology*; D021141:Nucleic Acid Amplification Techniques; D017924:Nucleopolyhedroviruses / Q000235:genetics / Q000472:pathogenicity*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014771:Virion / Q000235:genetics / Q000302:isolation & purification
D004279:DNA, Viral; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins