HeLaScribe® Nuclear Extract in vitro Transcription System

Cell-Free System for in vitro Transcription of Eukaryotic Genes

• Includes a CMV promoter positive-control template, rNTPs, Buffer, Stop Solution, Loading Dye and Nuclease-Free Water
• Performance-tested with cytomegalovirus immediate early gene (CMV) promoter
• Source of transcription factors, DNA-binding proteins and RNA processing components

The most well characterized cell-free system for in vitro transcription of eukaryotic genes is derived from HeLa cell nuclei. HeLa nuclear extracts can support accurate transcription initiation by RNA polymerase II and exhibit both basal and regulated patterns of RNA polymerase transcription. The nuclear extract is also a source of transcription factors, DNA-binding proteins and the enzymatic machinery involved in RNA processing.

The HeLa Nuclear Extract included in the HeLaScribe® Nuclear Extract in vitro Transcription System is prepared by a modification of the method of Dignam et al. Extracts prepared by this method have been shown to allow transcription from the human transferrin gene promoter and the adenovirus 2 major late promoter.

The system includes all the necessary components for in vitro transcription as well as a positive control template (CMV immediate early promoter DNA), and is performance-tested with the cytomegalovirus immediate early gene (CMV) promoter. HeLaScribe® Nuclear Extract is also available separately.

Applications of the HeLaScribe® Nuclear Extract in vitro Transcription System include:

• Analyzing basal and transcription factor-regulated transcription.
• Assessing Promoter strength.
• Mapping mRNA start sites.

References

1. Dignam, J.D. et al. (1983) Nucl. Acids Res. 11, 1475–89.
2. Sawadogo, M. and Roeder, R.G. (1985) Proc. Natl. Acad. Sci. USA 82, 4394–8.
3. Briggs, M.R. et al. (1986) Science 234, 47–52.
4. Bagchi, M.K. et al. (1990) J. Biol. Chem. 265, 5129–34.
5. Carthew, R.W. et al. (1985) Cell 43, 439–48.
6. Kristie, T.M. et al. (1989) EMBO J. 8, 4229–38.
7. Sawadogo, M. et al. (1988) J. Biol. Chem. 263, 11985–93.
8. Krainer, A.R. et al. (1984) Cell 36, 993–1005.

ItemPart #SizeConcentrationAvailable Separately

MgCl₂A357A1 × 500μl

50mMHeLaScribe® Nuclear Extract Transcription Grade

E309A2 × 20 reactionsView Product

HeLa Nuclear Extract 1X Transcription Buffer

E360A1 × 400μl

HeLa Extract Stop Solution

E361A1 × 15ml

HeLa Nuclear Extract Positive Control DNA

E362A1 × 300ngView Product

Loading Dye

E371A1 × 1mlrATP, 100mM

E601A1 × 15μlView Product

rUTP, 100mM

E602A1 × 15μlView Product

rGTP, 100mM

E603A1 × 15μlView Product

rCTP, 100mM

E604A1 × 15μlView Product

Nuclease-Free Water

P119A1 × 1,250μlView Product

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