T7 RNA polymerase 5000U

Name: T7 RNA polymerase 5000U
Brand: HYGIA
Price: 199.00 USD
Availability: InStock

Based on 0 reviews. - Write a review

$199.00

Ex Tax: $199.00

Stock: In Stock
Model: 0459

HYGIA

Qty

Add to Cart

Buy Now Ask Question

Add to Wish List Compare this Product

Description
Reviews

Product Description:	T7 RNA Polymerase is a DNA-dependent RNA polymerase present in E. coli infected with bacteriophage T7. The enzyme has extremely high specificity for promoter sequences found in T7 bacteriophage DNA and in various cloning vectors containing the T7 promoter. This enzyme requires double-stranded DNA that contains the T7 promoter sequence as a template and NTPs as substrates, and synthesizes single-stranded RNA complementary to the template downstream of the promoter. The use of this enzyme together with the reaction buffer included in this product can produce a large quantity of RNA with high quality.
Applications:	Synthesis of unlabeled and labeled RNA that can be used:l for hybridization, in vitro RNA translation.l as a RNA, siRNA , substrate in RNase protection assays, template for genomic DNA sequencing.l in studies of RNA secondary structure and RNA-protein interactions, RNA splicing.
Source:	Purified from E. coli expressing the phage T7 RNA Polymerase gene on a plasmid.
Purity:	≥95% by SDS-PAGE
Endogenous nucleic acid	＜1 pg/μL by qPCR
Concentration:	50 U/μL
Definition of Activity Unit:	One unit is defined as the amount of enzyme that incorporates 1 nmol of [ 3H]ATP into the acid-insoluble fraction in 1 hour at 37℃, pH 8.0.
5× Reaction Buffer:	200 mM Tris-HCl, 30 mM MgCl2, 50 mM DTT, 50 mM NaCl, 10 mM spermidine, pH 7.9
Storage (Dilution) Buffer:	50 mM Tris-HCl, 150 mM NaCl, 5 mM DTT, 0.1 mg/mL BSA, 50% Glycerol, pH 8.0
Inactivation or inhibition	l Inhibitors: metal chelators, enzyme activity is reduced by 50% at NaCl or KCl concentration above 150 mM.l Inactivated by heating at 70°C for 10 min or by addition of EDTA.
Storage Conditions:	Store at –20°C up to 12 months.

Assay Protocol / Procedures

Application Example (Protocol forin vitro transcription)

l Thaw frozen reagents, mix and centrifuge briefly.

l Keep enzymes and nucleotides on ice.

l Keep the Reaction Buffer at room temperature.

1. Prepare the following reaction mixture at room temperature:

Component	Volume
5×Reaction Buffer	4 μL
NTP Mixture	2 mM final concentration
Template DNA	0.5-1 μg
T7 RNA polymerase	1 μL
Nuclease-Free Water	To 20 μL
Total volume	20 μL

2. Incubate at 37°C for 2 hours.

3. Optional:AddRNase Inhibitor（recommendG3414）to1 U/μL final concentration toexclude contamination with RNases; AddInorganic Pyrophosphatase (Thermostable) (recommendG3421) to improve transcription yieldsignificantly.

4. Add2 μL0.5M ofEDTAor cool at-20℃ to stop the reaction.

Note

1. The transcription reaction should be performed under conditions that exclude contamination with RNases. The tips, tubes and water should be nuclease free. All the solutions should be made up in nuclease free water. Wearing gloves is advisable.

2. The reaction mixture can be scaled up or down.

3. To produce RNA transcripts of a defined length, plasmid DNA is linearized by restriction digestion downstream of the insert. Restriction enzymes which generate blunt ends or 5’-overhangs are preferred.

4. Spermidine contained in the buffer forms the complex with nucleic acids and may be precipitated as indissoluble materials. The template DNA should be added 2nd lastly in the reaction before enzymes.

5. Enzymesshould be placed on ice when used, and stored at -20℃ immediately after use. It is recommended to store separately.

6. For your safety and health, pleasewear safety glasses, gloves, or protective clothing.

For Research Use Only!