Insertional activation of a promoterless thymidine kinase gene

Journal Article · Mon Aug 01 04:00:00 EDT 1988 · Mol. Cell. Biol.; (United States)

DOI:https://doi.org/10.1128/MCB.8.8.3298· OSTI ID:6691932

Hiller, S; Hengstler, M; Kunze, M; Knippers, R

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), Journal Name: Mol. Cell. Biol.; (United States) Vol. 8:8; ISSN MCEBD
• Country of Publication: United States
• Language: English

Related Subjects

550200* -- Biochemistry
59 BASIC BIOLOGICAL SCIENCES
ALKALINE EARTH METAL COMPOUNDS
ANIMAL CELLS
AZINES
BIOCHEMICAL REACTION KINETICS
CALCIUM COMPOUNDS
CALCIUM PHOSPHATES
CELL CONSTITUENTS
CELL CULTURES
CLONING
CONNECTIVE TISSUE CELLS
DATA
DISEASES
DNA HYBRIDIZATION
DNA SEQUENCING
DNA-CLONING
ENZYMES
EXPERIMENTAL DATA
FIBROBLASTS
GENE OPERONS
GENE REGULATION
GENES
GENETIC ENGINEERING
HERPES SIMPLEX
HETEROCYCLIC COMPOUNDS
HYBRIDIZATION
INFECTIOUS DISEASES
INFORMATION
KINETICS
NUCLEOSIDES
NUCLEOTIDES
NUMERICAL DATA
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXYGEN COMPOUNDS
PHENOTYPE
PHOSPHATES
PHOSPHORUS COMPOUNDS
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
PLASMIDS
PRECIPITATION
PYRIMIDINES
REACTION KINETICS
RIBOSIDES
SEPARATION PROCESSES
SKIN DISEASES
SOMATIC CELLS
STRUCTURAL CHEMICAL ANALYSIS
THYMIDINE
TRANSFERASES
VIRAL DISEASES