Products

T7 RNA Polymerase

Bacteriophage T7 RNA Polymerase is a DNA-dependent RNA polymerase with high specificity for the T7 promoter. This enzyme catalyzes the 5’→3’synthesis of RNA from DNA downstream from its promoter.
No. Size Price Qty Status
C15010H-25000U 25,000 U (200 U/μL) $560.00 Inquiry
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Source:
Escherichia coli
 
Purity:
>98% as determined by SDS-PAGE. Purified by Ni-NTA chromatography.
 
Unit Definition:
One unit is defined as the amount of the enzyme incorporates 1 nmol of ATP into acid-insoluble product in 1 hour at 37°C.
 
Reaction Condition:
1X RNA Polymerase Reaction Buffer, supplemented with 0.5 mM each ATP, UTP, GTP, CTP, and DNA template containing the T7 RNA Polymerase promoter. Incubate at 37°C.
10X RNA Polymerase Reaction Buffer: 400 mM Tris-HCl (pH 8.0), 60 mM MgCl2, 100 mM DTT, and 20 mM spermidine.
 
Storage Buffer:
T7 RNA Polymerase is supplied in 100 mM Tris-HCl (pH 7.9), 20 mM KCl, 1 mM DTT, 1 mM EDTA, 0.1% Triton® X-100 and 50% (v/v) glycerol.

Storage:
Stored at -20°C. Avoid repeated freeze/thaw cycles.

Notes:
1. Transcription reaction should be performed under RNase free condition. Use nuclease-free tubes, reagents, and water to avoid RNase contamination. Also, wear gloves when working with RNA.
2. To obtain optimal condition, NTP concentration can be titrated between 10 – 15 mM.
3. The volume of T7 RNA Polymerase can be titrated between 1-2 μL in the IVT reaction to optimize your assay.
Standard RNA synthesis procedures:
1. Below reaction mixture should be prepared under room temperature and combined in the following order:
 
Component Amount Final concentration
Nuclease-Free H2O X μL -
Template DNA 0.5-1 μg -
10X RNA Polymerase Reaction Buffer 2 μL 1X
ATP (100 mM) 2 μL 10 mM
UTP (100 mM) 2 μL 10 mM
CTP (100 mM) 2 μL 10 mM
GTP (100 mM) 2 μL 10 mM
T7 RNA Polymerase (200 U/μL) 1 μL -
RNase inhibitor (optional) 0.5 μL 1U/μL
Total reaction volume 20 μL -
2. Incubate at 37°C for 30 minutes to 2 hours.
3. Above reaction mixture may be scaled up or down proportionately.