Rapid and efficient introduction of a foreign gene into bacterial artificial chromosome-cloned varicella vaccine by Tn7-mediated site-specific transposition.
Using a rapid and reliable system based on Tn7-mediated site-specific transposition, we have successfully constructed a recombinant Oka varicella vaccine (vOka) expressing the mumps virus (MuV) fusion protein (F). The backbone of the vector was our previously reported vOka-BAC (bacterial artificial chromosome) genome. We inserted the transposon Tn7 attachment sequence, LacZalpha-mini-attTn7, into the region between ORF12 and ORF13 to generate a vOka-BAC-Tn genome. The MuV-F expressing cassette was transposed into the vOka-BAC genome at the mini-attTn7 transposition site. MuV-F protein was expressed in recombinant virus, rvOka-F infected cells. In addition, the MuV-F protein was cleaved in the rvOka-F infected cells as in MuV-infected cells. The growth of rvOka-F was similar to that of the original recombinant vOka without the F gene. Thus, we show that Tn7-mediated transposition is an efficient method for introducing a foreign gene expression cassette into the vOka-BAC genome as a live virus vector.
Somboonthum
Pranee
P
Laboratory of Virology and Vaccinology, Division of Biomedical Research, National Institute of Biomedical Innovation, Japan.
Koshizuka
Tetsuo
T
Okamoto
Shigefumi
S
Matsuura
Masaaki
M
Gomi
Yasuyuki
Y
Takahashi
Michiaki
M
Yamanishi
Koichi
K
Mori
Yasuko
Y
D000818:Animals; D002460:Cell Line; D019433:Chickenpox Vaccine / Q000235:genetics*; D002522:Chlorocebus aethiops; D022202:Chromosomes, Artificial, Bacterial; D004251:DNA Transposable Elements; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors; D006168:Guinea Pigs; D016254:Mutagenesis, Insertional / Q000379:methods*; D011995:Recombination, Genetic; D014760:Viral Fusion Proteins / Q000235:genetics*; D019562:Viral Load; D010948:Viral Plaque Assay
D019433:Chickenpox Vaccine; D004251:DNA Transposable Elements; D014760:Viral Fusion Proteins