the PA-317 packaging line with a population of ty/ 2 cells transfected 48 hours 
previously with the SAX DNA. Of ten clones assayed because of high titer 
(>10^) and resistance to G418. one did not produce ADA by starch gel analysis 
nor would its supernatant confer ADA activity on the ADA(-) T lymphocytes 
(TJF-2). Using a Neo R probe. EcoRI digests of the genomic DNA from six ADA 
producing clones produced hybridization to the predicted 3.2 kb fragment. The 
ADA negative clone produced a fragment of 8.7 kb rather than the 3.2 kb 
expected of an intact vector. Presumably the loss of ADA function resulted 
from the deletion of a segment of the ADA gene including the EcoRI site. We 
have not determined if the SV40 promoter was left Intact. 
While the number of clones examined is too small to quantitate the 
statistical significance of these data, the results do suggest that the 
mutation rate during the initial transfection and Infection steps may be high. 
In summary, the SAX vector Is replication-defective so that theoretically 
only the cells exposed to the vector will be Infected. Helper-free systems 
should greatly reduce the spread of the retrovirus to other organs. Including 
the germ line, as well as to reduce the risk of malignancy by exposing the 
fewest possible cells to the mutagenic potential of retroviral insertion. 
II. Possibility for recombination leading to retrovirus production 
A concern with the use of retroviral vectors is the production of viral 
particles by cells receiving the vector. In a helper virus free system the 
production of viral particles could result by two possible mechanisms, both of 
which have different consequences. (1) Human endogenous retroviral sequences 
present in the host could recombine with the vector leading to a mutant virus 
that is then replication-competent. The new vector could establish a viremia 
Recombinant DNA Research, Volume 12 
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