Production of human alpha-interferon in silkworm using a baculovirus vector.
Microorganisms are generally used for mass production of foreign gene products, but multicellular organisms such as plants have been proposed as an economical alternative. The silkworm may be useful in this context as it can be cultured easily and at low cost. We have therefore developed a virus vector to introduce foreign genes, for example, the gene for human alpha-interferon (IFN-alpha), into silkworms. We used the baculovirus Bombyx mori nuclear polyhedrosis virus (BmNPV) which has a large (greater than 100 kilobases, kb) double-stranded circular DNA genome within its rod-shaped capsid. Baculoviruses have been used previously as vectors for expression of beta-interferon and beta-galactosidase in established cell lines. Although BmNPV has not been used previously as an expression vector, it has an advantage over the baculovirus Autographa californica NPV in that it has a narrower host range and will not grow in wild insect pests in the field. In the present study, the polyhedrin gene encoding the major inclusion body protein of BmNPV was identified by hybridization with complementary DNA and cloned in a plasmid. For insertion of foreign genes, we constructed a recombinant plasmid carrying a polylinker linked to the promoter of the polyhedrin gene, and inserted the IFN-alpha gene into this plasmid. The resulting plasmid and the BmNPV genomic DNA were co-transfected into BM-N cells, and stable recombinant viruses isolated by plaque assay on BM-N cells. The recombinant virus replicated in silkworm larvae, which synthesized as much as 5 X 10(7) units (approximately 50 micrograms) of interferon in their haemolymph.
Maeda
S
S
Kawai
T
T
Obinata
M
M
Fujiwara
H
H
Horiuchi
T
T
Saeki
Y
Y
Sato
Y
Y
Furusawa
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000235:genetics*; D004262:DNA Restriction Enzymes; D005822:Genetic Vectors; D006458:Hemolymph; D006801:Humans; D007304:Insect Viruses / Q000235:genetics*; D007370:Interferon Type I / Q000235:genetics* / Q000302:isolation & purification; D007700:Kinetics; D010957:Plasmids
D007370:Interferon Type I; D004262:DNA Restriction Enzymes