Overview 
Transfer and Expression of Cloned Genes Using 
Retroviral Vectors 
Eli Gilboa 1 , Martin A. Eglitis 2 , 
Philip W. Kantoff 2 and 
W. French Anderson 2 
'Dept, of Molecular Biology, 
Memorial Sloan-Kettering 
Cancer Center and 2 Laboratory 
of Molecular Hematology, 
National Heart, Lung and Blood 
Institute, NIH 
50-1 BioTcchniques 
INTRODUCTION 
It has been a little more than 10 years 
since the genetic engineering revolution 
began. Initially, "genetic engineering” 
applied only to manipulation of genes 
in bacteria, being a technology which 
enables the isolation and large scale pro- 
duction of specific DNA molecules 
from any source, whether or not they 
encode a gene. If properly engineered, 
such genetic manipulations can be used 
for the large scale production of specific 
gene products in bacteria and other sim- 
ple organisms. 
Genetic engineering of higher eucar- 
yotic or mammalian cells requires the 
introduction and expression of genes in 
these cells. One method for transferring 
genes into mammalian cells is DNA 
transfection in which DNA is intro- 
duced into cells in culture as part of a 
copredpitate with calcium phosphate or 
dextran sulfate (26). A successful result 
is a viable cell containing one to many 
copies of the new gene which contin- 
uously expresses the new genetic infor- 
mation. Unfortunately, DNA transfec- 
tion has its limitations. Most signifi- 
cantly, it is a very inefficient means of 
transferring genes into mammalian 
cells. At best only one in a thousand 
cells (more typically, one cell in a 
million) will incorporate the newly 
transferred gene. Additionally, not all 
cultured cell lines are susceptible to this 
method of gene transfer. Thus, a cen- 
tral issue in genetic engineering as ap- 
plied to mammalian cells is gene trans- 
fer: How to deliver a gene into a large 
fraction of any given cell population. A 
second important issue is the question 
of expression — once delivered, how to 
ensure the proper expression of the gene 
in the recipient cell. 
In the past several years, a new gene 
transfer technology has emerged which 
appears to be superior to the DNA 
transfection and other previous tech- 
niques and which may offer a new ap- 
proach to the therapy of human genetic 
diseases (1). This new technology is 
known as retroviral-mediated gene 
transfer, i.e., the use of retroviruses to 
deliver genes into cells. 
In order to explain how retroviruses 
can be utilized for gene transfer, it is im- 
portant to describe some of the salient 
features of their replication (For more 
detailed information on retroviruses, 
see Ref. 25.). Retroviruses are RNA 
viruses, that is, the viral genes are en- 
coded in an RNA molecule rather than 
in a DNA molecule. When the virus 
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Recombinant DNA Research, Volume 12 
