Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei.
نویسندگان
چکیده
We have developed a procedure for preparing extracts from nuclei of human tissue culture cells that directs accurate transcription initiation in vitro from class II promoters. Conditions of extraction and assay have been optimized for maximum activity using the major late promoter of adenovirus 2. The extract also directs accurate transcription initiation from other adenovirus promoters and cellular promoters. The extract also directs accurate transcription initiation from class III promoters (tRNA and Ad 2 VA).
منابع مشابه
A mammalian RNA polymerase II holoenzyme containing all components required for promoter-specific transcription initiation
The protein kinase MO15/CDK7 has recently been shown to be associated with the general transcription factor TFIIH and to be capable of phosphorylating the RNA polymerase II carboxy-terminal domain. Here, we show that a monoclonal MO15/CDK7 antibody coimmunoprecipitates, from a rat liver nuclear extract, all components of the RNA polymerase II transcription apparatus required for initiation at t...
متن کاملMapping of RNA polymerase on mammalian genes in cells and nuclei.
The assembly of an RNA polymerase II initiation complex at a promoter is associated with the melting of the DNA template to allow the polymerase to read the DNA sequence and synthesize the corresponding RNA. Using the specific single-stranded modifying reagent KMnO4 and a new genomic sequencing technique, we have explored the melted regions of specific genes in genomic DNA of whole cells or of ...
متن کاملIsolation of the protein and RNA content of active sites of transcription from mammalian cells.
Mammalian cell nuclei contain three RNA polymerases (RNAP I, RNAP II and RNAP III), which transcribe different gene subsets, and whose active forms are contained in supramolecular complexes known as 'transcription factories.' These complexes are difficult to isolate because they are embedded in the 3D structure of the nucleus. Factories exchange components with the soluble nucleoplasmic pool ov...
متن کاملFunctions of the N- and C-terminal domains of human RAP74 in transcriptional initiation, elongation, and recycling of RNA polymerase II.
Transcription factor IIF (TFIIF) cooperates with RNA polymerase II (pol II) during multiple stages of the transcription cycle including preinitiation complex assembly, initiation, elongation, and possibly termination and recycling. Human TFIIF appears to be an alpha2beta2 heterotetramer of RNA polymerase II-associating protein 74- and 30-kDa subunits (RAP74 and RAP30). From inspection of its 51...
متن کاملSterile preparation of antibiotic-selective LB agar plates using a microwave oven.
WI-L2-NS cells, a p53 mutant cell line. Mutat. Res. 352:79-86. 2.Cherbonnel-Lasserre, C., S. Gauny, and A. Kronenberg. 1996. Suppression of apoptosis by Bcl-2 or Bcl-xL promotes susceptibility to mutagenesis. Oncogene 13:1489-1497. 3.Dignam, J.D., R.M. Lebovitz, and R.G. Roeder. 1983. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nuc...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
- Nucleic acids research
دوره 11 5 شماره
صفحات -
تاریخ انتشار 1983