An Assay for Transient Gene Expression in Transfected Drosophila Cells, Using [3H]guanine Incorporation

Overview

Journal EMBO J

Specialties Biotechnology
Molecular Biology

Date 1984 Nov 1

PMID 6096129

Citations 13

Authors

J F Burke

J H Sinclair

J H Sang

D Ish-Horowicz

Affiliations

Soon will be listed here.

Abstract

We have developed an assay for transient gene expression using a dominant-selectable marker previously employed to transform Drosophila cultured cells. Drosophila hydei cells transfected with a functional Escherichia coli xanthine guanine phosphoribosyl transferase gene (gpt), under the control of the long terminal repeats (LTRs) of the copia transposable element, rapidly incorporate guanine into acid-precipitable counts. Autoradiographic analysis in situ shows that approximately 20% of cells take up, and express, the gpt gene. This transient gpt expression depends on the Drosophila promoter sequences since vectors with the gpt gene in reverse orientation to the copia LTRs fail to incorporate guanine. Deletion analysis confirms that the LTRs are essential for gpt gene expression. Similarly, cells transfected with gpt controlled by the Drosophila 70 000 mol. wt. heat-shock (hsp 70) promoter show regulated guanine incorporation when heat shocked. The efficiency of the copia LTRs varies considerably between the cell lines we tested, whereas that of the hsp 70 promoter does not. The heterologous promoters of the Rous sarcoma virus (RSV) and simian virus 40 (SV40) function poorly in these cells.

Citing Articles

Functional analysis of the transcriptional control regions of the copia transposable element.

Sinclair J, Burke J, Ish-Horowicz D, Sang J EMBO J. 1986; 5(9):2349-2354.

PMID: 16453706 PMC: 1167119. DOI: 10.1002/j.1460-2075.1986.tb04503.x.

Hormonal and developmental specificities of transcription lie within a 1.5 kb region 5' to the ecdysone inducible Drosophila P1 gene.

Maschat F, Roux J, Benes H, Pictet R, Jami J, Lepesant J EMBO J. 1986; 5(3):583-8.

PMID: 16453674 PMC: 1166802. DOI: 10.1002/j.1460-2075.1986.tb04250.x.

Regulated expression of a Drosophila melanogaster heat shock locus after stable integration in a Drosophila hydei cell line.

Sinclair J, Saunders S, Burke J, Sang J Mol Cell Biol. 1985; 5(11):3208-13.

PMID: 3939313 PMC: 369136. DOI: 10.1128/mcb.5.11.3208-3213.1985.

20-Hydroxyecdysone increases levels of transient gene expression in transfected Drosophila cells.

Sinclair J, Bryant L Nucleic Acids Res. 1987; 15(22):9255-61.

PMID: 3120150 PMC: 306466. DOI: 10.1093/nar/15.22.9255.

Transformation of cultured Drosophila melanogaster cells with a dominant selectable marker.

Rio D, Rubin G Mol Cell Biol. 1985; 5(8):1833-8.

PMID: 3018529 PMC: 366898. DOI: 10.1128/mcb.5.8.1833-1838.1985.

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