DEVELOPMENT OF AVIAN-SARCOMA AND LEUKOSIS VIRUS-BASED VECTOR-PACKAGING CELL-LINES

We have constructed an avian leukosis virus derivative with a 5'' deletion extending from within the tRNA primer binding site to a SacI site in the leader region. Our aim was to remove cis-acting replicative and/or encapsidation sequences and to use this derivative, RAV-1.PSI.-, to develop vector-packaging cell lines. We show that RAV-1.PSI.- can be stably expressed in the quail cell line QT6 and chicken embryo fibroblasts and that it is completely replication deficient in both cell types. Moreover, we have demonstrated that QT6-derived lines expressing RAV-1.PSI.- can efficiently package four structurally different replication-defective v-src expression vectors into infectious virus, with very low or undetectable helper virus release. These RAV-1.PSI.--expressing cell lines comprise the first prototype avian sarcoma and leukosis virus-based vector-packaging system. The construction of our vectors has also shown us that a sequence present within gag, though to facilitate virus packaging, is not necessary for efficient vector expression and high virus production. We show that quantitation and characterization of replication-defective viruses can be achieved with a sensitive immunocytochemical procedure, presenting an alternative to internal selectable vector markers.