The HeLa cell nuclear extract, catalog number P0001, is specifically recommended for: 1) in vitro transcription, 2) protein-DNA/RNA and protein-protein interactions, and 3) as a source of individual transcription factors. Although this extract also contains most pre-mRNA processing factors, it is, however, not recommended for in vitro pre-mRNA processing.

The HeLa cell nuclear extract was prepared as described by Dignam et al. (1) and Manley et al. (2). This extract contains all basal transcription factors, including TFIIA, -IIB, -IID, -IIE, -IIF, -IIH and RNA polymerase II, as well as most gene-specific activators and cofactors, such as Sp1, Oct-1, NF-kB, USF, ATF, PC4, p300 etc. (3-7). Therefore, it has been used as the source of transcription factors for the cell-free transcription system to study specific transcription by RNA polymerase II as well as RNA polymerase III (8) and for purifying transcription factors (9, 10).

Source
Extracted from Cervical Tumor cells

Concentration
8 mg protein/ml (in 1x dilution buffer A)

Unit Definition (Activity)
1-5 μl is sufficient for a gel mobility shift assay in a 20 μl reaction; 5-10 μl is sufficient for in vitro transcription assay and 20-50 μl is sufficient for a protein-protein interaction assay. Extract activity has been validated using an in vitro transcription assay.

Buffer and Storage
1X dilution buffer A: 20 mM Tris-Cl (pH 8.0), 20% Glycerol, 100 mM KCl, 1 mM DTT and 0.2 mM EDTA. The extract is stored at -80ºC. Avoid repeated freezing and thawing cycles.

Applications
The HeLa cell nuclear extract of this catalog number is specifically recommended for: 1) in vitro transcription; 2) protein-DNA/RNA and protein-protein interactions; and 3) as a source of individual transcription factors. Although this extract also contains most pre-mRNA processing factors, it is, however, not recommended for in vitro pre-mRNA processing.

References
1. Dignam, J.D., et al., (1983) Nucleic Acids Res. 11, 1475-1489
2. Manley, J.L., et al., (1980) Proc. Natl. Acad. Sci. USA 77, 5706-5710
3. Sawadogo, M., et al., (1985) Proc. Natl. Acad. Sci. USA 82, 4394-4398
4. Kadonaga, J.T., et al., (1986) Proc. Natl. Acad. Sci. USA 83, 5889-5893
5. Reinberg, D., et al., (1987) J. Biol. Chem. 262, 3322-3330
6. Horikoshi, M., et al., (1988) Cell 54, 1033-1042
7. Yang, X.J., et al., (1996) Nature 382, 19-24
8. Hoeffler, W.K., et al., (1988) Cell 53, 907-920
9. Ge, H., et al., (1996) Methods Enzymol. 274, 57-71
10. Maldonado, E., et al., (1996) Methods Enzymol. 274, 72-100

DISCLAIMER

This products is recommended For RESEARCH USE ONLY and is Not qualified for Use in Diagnostic or Therapeutic Procedures.