THE EFFECT OF SELECTION FOR HIGH-LEVEL VECTOR EXPRESSION ON THE GENETIC AND FUNCTIONAL STABILITY OF A SINGLE TRANSCRIPT VECTOR DERIVED FROMA LOW-LEUKEMOGENIC MURINE RETROVIRUS

Single-gene murine leukemia virus-based retroviral vectors carrying th e G418-resistance gene (neo) under transcriptional control of the long terminal repeat were used to study the effect of selection on long-te rm vector expression in a murine lymphoid cell line, L691. We used two isogenic vectors carrying either a strong or a weak transcriptional e nhancer from low-leukemogenic Akv and high-leukemigenic SL3-3 murine l eukemia virus, respectively. Effects of G418 selection were studied at the level of vector-transduced cell populations and at the level of s ingle-vector-transduced cell clones obtained without selection for vec tor expression. Selection for vector expression prior to isolation of cell clones changed the range of vector expression levels for cell clo nes carrying the Akv enhancer, but did not influence the long-term sta bility of vector expression for the two populations of cell clones. Ce ll clones harboring the Akv enhancer, isolated without selection and t hen subjected to prolonged growth under selective conditions, exhibite d no mutations in the enhancer region or major vector rearrangements a lthough showing increased vector expression in some cases. Our results are discussed in terms of retrovirus-mediated gene transfer strategie s employing selection for expression of a selective marker in single-g ene or bicistronic vectors with a low- or nonleukemogenic virus-derive d backbone.