(b) Infection. After the producer cell line Is established, supernatant 
(i.e., RVS) can be collected and the target cells of choice can be infected. 
The mutation rate of retroviral genomes after infection is high. Shields et 
al . (1978) Infected 3T3 cells with Moloney MuLV and found that 1/3 of the 
single virus cell clones expressed viral gene function aberrantly. Goff et al . 
(1981) studied wild type Moloney MuLV and recovered temperature sensitive 
mutants with a frequency of 3-5%. 
Gene transfer by transfection or infection can lead to loss of certain 
regions within the viral genome. Joyner and Bernstein (1983) constructed a 
vector derived from spleen focus-forming virus (SFFV) which contained the 
thymidine kinase (TK) gene within its noncoding region. When infectious 
particles that conferred TK expression were selected they were no longer able 
to cause erythrol eukemia. When the same vector was selected for its ability to 
produce erythrol eukemia. It no longer expressed the TK gene. Of significance 
to our work is the finding that vectors containing internal promoters (such as 
our SV40) may be associated with a high frequency of deletions and that the 
deleted vectors may have a selective advantage since they appear to have 
increased multiplicity and titer (Bandyopadhyay and Temin, 1984; Emerman and 
Temin, 1984). 
(2) Stability of the inserted vector genome. 
The stability of the SV40 internal promoter used in the SAX vector has not 
been studied definitively. We have shown that the SAX vector can be stably 
transfected and infected into a variety of in vitro cell lines under selection 
but have not systematically assessed the percentage of stable, intact 
insertions versus defective integration. 
An example of the type of rearrangements that can occur during the initial 
transfection is the following. We have assayed clones obtained by infection of 
[52] 
Recombinant DNA Research, Volume 12 
