Insertional activation of a promoterless thymidine kinase gene
A plasmid carrying a promoterless herpes complex virus thymidine kinase gene was transfected via calcium phosphate precipitation into LM (tk/sup -/) mouse fibroblast cells. The transfected gene was efficiently expressed, as the transfected cells grew perfectly well in selective hypoxanthine-aminopterin-thymidine medium, suggesting that the thymidine kinase-coding region became linked to a promoterlike element on integration into the recipient genome. To investigate the structure of the surrogate promoter, the authors first isolated the integrated gene from a genomic library. The nucleotide sequence of the DNA adjacent to the thymidine kinase-coding sequence was then determined. They found, first, that the integration of the transfected DNA apparently occurred by a blunt end ligation mechanism involving no obvious sequence similarities between integrated and recipient DNA and, second, that the 5'-flanking region included a TATA box, to CCAAT boxes, and a GC box element. However, the TATA box motif and the most proximal CCAAT box appeared to be sufficient of full promoter activity, as determined by the transfection efficiencies of appropriate plasmid constructs. Except for these canonical promoter elements, the surrogate promoter had no obvious similarities to known thymidine kinase gene promoters.
- Research Organization:
- Deutsches Krebsforschungszentrum, Heidelberg (DE)
- OSTI ID:
- 6691932
- Journal Information:
- Mol. Cell. Biol.; (United States), Vol. 8:8
- Country of Publication:
- United States
- Language:
- English
Similar Records
Transfer of nonselectable genes into mouse teratocarcinoma cells and transcription of the transferred human. beta. -globin gene
Phenotypic switching in cells transformed with the herpes simplex virus thymidine kinase gene
Related Subjects
GENE OPERONS
DNA SEQUENCING
GENE REGULATION
HERPES SIMPLEX
BIOCHEMICAL REACTION KINETICS
PHOSPHOTRANSFERASES
DNA-CLONING
CALCIUM PHOSPHATES
CELL CULTURES
EXPERIMENTAL DATA
FIBROBLASTS
GENES
GENETIC ENGINEERING
PHENOTYPE
PLASMIDS
PRECIPITATION
THYMIDINE
ALKALINE EARTH METAL COMPOUNDS
ANIMAL CELLS
AZINES
CALCIUM COMPOUNDS
CELL CONSTITUENTS
CLONING
CONNECTIVE TISSUE CELLS
DATA
DISEASES
DNA HYBRIDIZATION
ENZYMES
HETEROCYCLIC COMPOUNDS
HYBRIDIZATION
INFECTIOUS DISEASES
INFORMATION
KINETICS
NUCLEOSIDES
NUCLEOTIDES
NUMERICAL DATA
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXYGEN COMPOUNDS
PHOSPHATES
PHOSPHORUS COMPOUNDS
PHOSPHORUS-GROUP TRANSFERASES
PYRIMIDINES
REACTION KINETICS
RIBOSIDES
SEPARATION PROCESSES
SKIN DISEASES
SOMATIC CELLS
STRUCTURAL CHEMICAL ANALYSIS
TRANSFERASES
VIRAL DISEASES
550200* - Biochemistry