N1648
HeLa cell nuclear extract
usage
vial sufficient for 50 reactions
shipped in
dry ice
storage temp.
−70°C
Application
A cell-free transcription system that produces accurate transcripts with RNA polymeraseII. Reaction will incorporate 50-150fmol nucleotides into a 400-nucleotide transcript from Ad2ML (adenovirus major late promoter) in 60min at 30°C.
Biochem/physiol Actions
The system carries out basal transcription, dependent on recognition of the TATA element, as well as transcription which is regulated by interaction of transcription factors with upstream promoter elements, e.g., USF, Oct-1, Sp-1, progesterone receptor.
Depleting the extract of individual transcription factors produces a system for assaying purified factors (TATA binding protein, Oct-1, etc.) by complementation. The extract is also a source of transcriptional regulators and RNA processing proteins.
Depleting the extract of individual transcription factors produces a system for assaying purified factors (TATA binding protein, Oct-1, etc.) by complementation. The extract is also a source of transcriptional regulators and RNA processing proteins.
Packaging
Supplied in 20 mM HEPES, pH 9.0, containing 100 mM KCl, 0.2 mM EDTA, 1 mM DTT, and 20% glycerol.
Analysis Note
The extract has been treated with PMSF to inhibit proteases
Regulatory Information
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J L Manley et al.
Proceedings of the National Academy of Sciences of the United States of America, 77(7), 3855-3859 (1980-07-01)
We have developed a cell-free system for studying the synthesis of mRNA in mammalian cells. The system consists of a dialyzed and concentrated whole-cell extract derived from HeLa cells, small molecules and cofactors needed for transcription, and exogenously added DNA.
Nakajima, N., et al.
Molecular and Cellular Biochemistry, 8, 4028-4028 (1988)
Regulation of in vitro transcription by progesterone receptor. Characterization and kinetic studies.
M K Bagchi et al.
The Journal of biological chemistry, 265(9), 5129-5134 (1990-03-25)
We have devised an in vitro assay system to study the transcriptional activity of native chicken progesterone receptor (cPR). Purified cPR added to cell-free extracts from HeLa cell nuclei stimulates accurate transcription from a promoter driven by two progesterone response
T M Kristie et al.
The EMBO journal, 8(13), 4229-4238 (1989-12-20)
The herpes simplex virus transactivator, alpha TIF, stimulates transcription of the alpha/immediate early genes via a cis-acting site containing an octamer element and a conserved flanking sequence. The alpha TIF protein, produced in a baculovirus expression system, nucleates the formation
Sawadogo, M., et al.
The Journal of Biological Chemistry, 263, 11985-11985 (1985)
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