A simple method for the simultaneous detection of E1A and E1B in adenovirus stocks.
Recombinant adenoviral vectors have been developed for use as therapeutic agents and for the introduction of exogenous genes into living cells. However, the occurrence of replication-competent adenoviruses (RCA) in adenovirus stocks produced in 293 cells remains a major problem in terms of the safe use of such vectors. To overcome the problems associated with the occurrence of RCA, we have established a simple method for the simultaneous detection of amplified E1A and E1B from RCA that might contaminate adenoviral stocks. The products amplified by polymerase chain reaction (PCR) were fractionated by regular electrophoresis on agarose gels and visualized by staining with ethidium bromide. This method is rapid and inexpensive for detection of RCA in the preparation of adenoviruses.
Suzuki
Erika
E
Gene Enginering Division, Division of Biological Systems, BioResource Center, Tsukuba Institute, RIKEN (The Institute of Physical and Chemical Research), 3-1-1 Koyadai, Tsukuba Science City, Ibaraki 305-0074, Japan.
Murata
Takehide
T
Watanabe
Sanae
S
Kujime
Yukari
Y
Hirose
Megumi
M
Pan
Jianzhi
J
Yamazaki
Takahito
T
Ugai
Hideyo
H
Yokoyama
Kazunari K
KK
D000256:Adenoviridae / Q000145:classification / Q000235:genetics*; D017500:Adenovirus E1A Proteins / Q000235:genetics*; D017501:Adenovirus E1B Proteins / Q000235:genetics*; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D006367:HeLa Cells; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011995:Recombination, Genetic; D014779:Virus Replication
D017500:Adenovirus E1A Proteins; D017501:Adenovirus E1B Proteins; D017931:DNA Primers; D004279:DNA, Viral