Adaptation of a retrovirus as a eucaryotic vector transmitting the herpes simplex virus thymidine kinase gene
The authors investigated the feasibility of using retroviruses as vectors for transferring DNA sequences into animal cells. The thymidine kinase (tk) gene of herpes simplex virus was chosen as a convenient model. The internal BamHI fragments of a DNA clone of Moloney leukemia virus (MLV) were replaced with a purified BamHI DNA segment containing the tk gene. Chimeric genomes were created carrying the tk insert on both orientations relative to the MLV sequence. Each was transfected into TK/sup -/ cells along with MLV helper virus, and TK/sup +/ colonies were obtained by selection in the presence of hypoxanthine, aminopterin, and thymidine (HAT). Virus collected from TK/sup +/-transformed, MLV producer cells passed the TK/sup +/ phenotype to TK/sup -/ cells. Nonproducer cells were isolated, and TK/sup +/ transducing virus was subsequently rescued from them. The chimeric virus showed single-hit kinetics in infections. Virion and cellular RNA and cellular DNA from infected cells were all shown to contain sequences which hybridized to both MLV- and tk-specific probes. The sizes of these sequences were consistent with those predicted for the chimeric virus. In all respects studied, the chimeric MLV-tk virus behaved like known replication-defective retroviruses. These experiments suggest great general applicability of retroviruses as eucaryotic vectors.
- Research Organization:
- Center for Cancer Research and Dept. of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139
- OSTI ID:
- 5963927
- Journal Information:
- Mol. Cell. Biol.; (United States), Vol. 2:4
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
HERPES SIMPLEX
ETIOLOGY
MOLECULAR BIOLOGY
STRUCTURE-ACTIVITY RELATIONSHIPS
PHOSPHOTRANSFERASES
DNA-CLONING
ENZYME ACTIVITY
VIRUSES
BIOLOGICAL ADAPTATION
ANIMAL CELLS
BIOLOGICAL MODELS
CELL CULTURES
CELL TRANSFORMATIONS
CHIMERAS
DNA SEQUENCING
EXPERIMENT PLANNING
GENES
GENOME MUTATIONS
HYBRIDIZATION
HYPOXANTHINE
LEUKEMIA VIRUSES
RIBOSOMAL RNA
STRUCTURAL MODELS
THYMIDINE
VIRULENCE
AROMATICS
AZAARENES
AZINES
CLONING
DISEASES
ENZYMES
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
INFECTIOUS DISEASES
MICROORGANISMS
MOSAICISM
MUTATIONS
NUCLEIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ONCOGENIC VIRUSES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PARASITES
PHOSPHORUS-GROUP TRANSFERASES
PLANNING
PURINES
PYRIMIDINES
RIBOSIDES
RNA
SKIN DISEASES
STRUCTURAL CHEMICAL ANALYSIS
TRANSFERASES
VIRAL DISEASES
550400* - Genetics
550300 - Cytology
550200 - Biochemistry