Rapid and efficient introduction of a foreign gene into bacterial artificial chromosome-cloned varicella vaccine by Tn7-mediated site-specific transposition

Abstract 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, LacZα–mini- att Tn7, 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- att Tn7 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.