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  • E. coli RNA Polymerase, Core Enzyme

    Description

    E. coli RNA Polymerase, Core Enzyme consists of 5 subunits designated α, α, β´, β, and ω. The enzyme is free of sigma factor and does not recognize any specific bacterial or phage DNA promoters. The enzyme retains the ability to transcribe RNA from nonspecific initiation sequences. Addition of sigma factors will allow the enzyme to initiate RNA synthesis from specific bacterial and phage promoters. The core enzyme has a molecular weight of approximately 400 kDa.

    Product Source

    E. coli RNA Polymerase, Core Enzyme is isolated from E. coli strain BL21.

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    E.coli RNA Polymerase Reaction Buffer5X

    Advantages and Features

    Applications

    • RNA synthesis from E. coli promoter
    • Transcription initiation studies
    • In vitro translation with PURExpress

    Properties and Usage

    Unit Definition

    One unit is defined as the amount of enzyme required to incorporate 1 nmol NTP into RNA in 10 minutes at 37°C.

    Storage Temperature

    -20°C

    Storage Conditions

    20 mM Tris-HCl
    100 mM NaCl
    0.1 mM EDTA
    1 mM DTT
    50% Glycerol
    pH 7.5 @ 25°C

    Unit Assay Conditions

    1X E. coli RNA Polymerase Reaction Buffer, supplemented with 0.5 mM of each NTP, sigma factor 70 and 1 μg T7 phage DNA in 50 μl.

    Quality Control

    Quality Assurance Statement

    • E. coli RNA Polymerase, Core Enzyme is free of detectable DNA endonuclease, exonuclease and RNase activities.

    Quality Control Assays

    The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page.
    • Endonuclease Activity (Nicking):
      The product is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
    • Exonuclease Activity (Radioactivity Release):
      The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activity is determined by the % release of radioactive nucleotides.
    • RNase Activity (2 Hour Digestion):
      The product is tested in a reaction containing a RNA substrate.  After incubation for 2 hours there is no detectable degradation of the RNA substrate as determined by gel electrophoresis.

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.

    Datacards

    The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.
    1. What is the difference between the E.coli RNA Polymerase, Core Enzyme and Holoenzyme?
    2. What are the major functions of these subunits?
    3. Can the Core and Holoenzyme be used in PURExpress?
    4. What are the other sigma factors in E.coli?