[Design and construction of transfer vectors in order to obtain recombinant modified vaccinia virus Ankara (MVA)]

Modified Vaccinia virus Ankara (MVA) constitutes a good candidate for the development of non-replicative expression viral vectors because it does not replicate in most of mammalian cells. It is essential, for the production of recombinant MVA, the availability of transfer vectors which allow the introduction of desired genes into non-essential regions for in vitro viral replication, by homologous recombination with the viral genome. In the present work, the transfer vectors named VT-MHA and VT-MTK were designed and obtained. They carried genomic regions corresponding to 1-303 and 608-948 positions of the MVA165R gene and 1-244 and 325-534 of the MVA086R gene, respectively, which flank a multiple cloning site for the insertion of foreign genes. In these vectors, the cassettes for the expression of lac Z or uidA genes were cloned, and the activity of the marker enzymes beta-galactosidase and beta-glucuronidase was confirmed in situ. Furthermore, the vector named VT-MTK-GUS was used to obtain and isolate pure recombinant MVA, which carried and expressed the uid A gene. The results herein constitute the basic tools for establishing the methodology to obtain recombinant MVA with the purpose of locally developing non-replicative viral vectors as candidate vaccines.