The SAX retroviral vector has been used by our laboratories to efficiently 
transfer the Neo R and human ADA genes into transformed ADA-deficient T and B 
cells. Cells expressing the Neo R gene can be selected in G418-containing 
medium which also serves to enrich for cells expressing the introduced ADA 
gene. The SAX-transduced cells produced the appropriate human ADA isozymes 
when analysed by starch gel electrophoresis indicating correct post-trans- 
lational modification of a single gene product. This was also shown to be true 
when NIH 3T3 cells and a monkey kidney line (CV-1 cells) were infected with the 
SAX vector (McLachlin et al.. 1987). Quantitation of the levels of ADA 
enzymatic activity showed that the ADA levels in SAX-transduced ADA-deficient T 
and B cells exceeded the ADA production in normal, control cell lines. None of 
these changes (except growth in G418) were observed when ADA(-) cell lines were 
Infected with the parent vector, N2. which does not contain any human ADA gene 
sequences. Kantoff et al.. (1986) demonstrated that the ADA enzyme produced 
by the SAX-Infected cells was sufficient to dramatically reduce the sensitivity 
of these cells to high levels of 2 '-deoxyadenoslne indicating the functional 
correction of the ADA deficiency. 
D 
A similar retroviral vector. pZ IP-SV (B) ADA. containing the Neo gene and 
a human ADA cDNA, has been used by Belmont et al . (1986) for gene transfer into 
D 
murine bone marrow Ini vitro . Expression of the Neo gene was demonstrated for 
approximately 10% of the cells by colony formation in the presence of G418. 
Marrow cells which had been infected and selected in G418 were pooled to 
successfully demonstrate the production of human ADA enzyme by isoelectric 
focusing. Some CFU-C were also selected for Increased ADA expression using 
medium containing a combination of 9-xyl ofuranosyl adenine (XylA) and 
deoxycoformycin. a potent Inhibitor of ADA. 
Recombinant DNA Research, Volume 12 
[ 39 ] 
