Efficient transformation of primary human amniocytes by E1 functions of Ad5: generation of new cell lines for adenoviral vector production.
Primary human cells are relatively refractory to transformation by adenoviral E1 functions. For almost two decades, human embryonic kidney (HEK)-derived 293 cells have been the only E1-complementing cell line suitable for production of E1-deleted adenoviral vectors. More recently, new vector production cell lines have been derived from human embryonic retina (HER) cells, a cell type that is difficult to obtain. We were surprised to find that readily available primary human amniocytes are efficiently transformed by adenoviral E1 functions. We selected cell lines that allow high-titer production of recombinant adenoviral vectors. The generation of replication-competent adenovirus (RCA) during production, caused by homologous recombination between vector and cellular DNA, was excluded by designing the transforming plasmid to lack sequence overlap with current adenoviral vectors. In addition, we generated an infectious plasmid that can be used for convenient generation of first-generation adenoviral vectors in Escherichia coli and that matches the E1 complementation in the new production cell lines.
Schiedner
G
G
Center for Molecular Medicine (ZMMK), University of Cologne, 50931 Cologne, Germany. Gudrun.Schneider@medizin.uni-koeln.de
Hertel
S
S
Kochanek
S
S
D000256:Adenoviridae / Q000235:genetics*; D017498:Adenovirus E1 Proteins / Q000235:genetics*; D000653:Amniotic Fluid / Q000166:cytology*; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D008957:Models, Genetic; D010957:Plasmids / Q000378:metabolism; D011995:Recombination, Genetic; D013997:Time Factors; D014162:Transfection; D014170:Transformation, Genetic
D017498:Adenovirus E1 Proteins