Regulatory elements in the introns of the human HPRT gene are necessary for its expression in embryonic stem cells

Journal Article · Fri Jun 01 04:00:00 EDT 1990 · Proceedings of the National Academy of Sciences of the United States of America; (United States)

DOI:https://doi.org/10.1073/pnas.87.11.4299· OSTI ID:5200965

Reid, L H; Smithies, O; Koller, B H ^[1]; Gregg, R G ^[2]

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1. Univ. of North Carolina, Chapel Hill (USA)
2. Univ. of Wisconsin, Madison (USA)

The authors have examined the expression of transfected human hypoxanthine phosphoribosyltransferase minigenes (HPRT) in mouse embryonic stem (ES) cells. cDNA constructs of this gene that have been successfully used in somatic cell lines failed to confer hypoxanthine/aminopterin/thymidine (HAT) resistance in ES cells. In contrast, constructs containing introns 1 and 2 from the HPRT gene produced a high frequency of HAT-resistant colonies. This observation allowed them to identify two sequences in these introns that influence expression of the HPRT gene in ES cells. One element, located in intron 2, is required for effective HPRT expression in thee cells; the other element, located in intron 1, acts as an enhancer of HPRT expression. Using this information, they have constructed an HPRT minigene that can be used for either positive or negative selection in ES cell experiments. This dual capability allows the design of in-out procedures to create subtle changes in target genes by homologous recombination with the aid of this selectable minigene.

• OSTI ID: 5200965
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America; (United States), Journal Name: Proceedings of the National Academy of Sciences of the United States of America; (United States) Vol. 87:11; ISSN 0027-8424; ISSN PNASA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
CELL CONSTITUENTS
DNA
DNA SEQUENCING
EMBRYONIC CELLS
ENZYMES
GENE REGULATION
GLYCOSYL TRANSFERASES
HYPOXANTHINE PHOSPHORIBOSYLTRANSFERASE
MAMMALS
MAN
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PENTOSYL TRANSFERASES
PLASMIDS
PRIMATES
RECOMBINANT DNA
STRUCTURAL CHEMICAL ANALYSIS
TRANSFERASES
VERTEBRATES