T7 RNA Polymerase

Description

Bacteriophage T7 RNA Polymerase (RNAP) is a highly processive DNA-dependent RNA polymerase that catalyzes the transcription of T7 genes¹. It acts by unwinding the double-stranded DNA to provide access to the coding strand for transcription initiation (Figure 1). The 99 kDa enzyme catalyzes in vitro RNA synthesis from a cloned DNA sequence under T7 promoters. RNA produced using the T7 RNAP is suitable for many applications in research and biotechnology.

Intact Genomics IG^® T7 RNAP is a purified recombinant T7 bacteriophage enzyme that is expressed and purified from E. coli.  This enzyme is designed to perform in vitro transcription of a gene under a phage T7 promoter. The physical purity of this enzyme is ≥85% as assessed by SDS-PAGE with Coomassie^® blue staining and gel densitometry (Figure 2).

Fig. 1: The T7 RNAP enzyme is a protein that binds to and transcribes RNA from double-stranded DNA. A molecular surface representation of T7 RNAP in complex with DNA (PDB ID# 3e2e) is shown in the early stages of transcription. Protein residues-green, DNA-orange and blue strands, Initiating RNA strand-black.

Applications

• Radiolabeled RNA probe preparation
• RNA synthesis from DNA template
• sgRNA and mRNA synthesis
• Applications in mRNA therapeutics

Product Source

E. coli BL21 (DE3) strain expressing a recombinant T7 RNA polymerase gene (T7 gene 1) with an N-terminal 6xHis tag.

Components and Storage

The T7 RNA Polymerase product contains the items below.  Store all components at -20°C.

• T7 RNA Polymerase 25000 Units (50,000 Units/mL)
• Storage Buffer (50 mM Tris-HCl, 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 50% Glycerol, pH 7.5 @ 25 ºC)

Quality Control Assays

T7 RNA Polymerase is free from detectable RNase, Endonuclease (nicking), and DNase activities.

Functional Testing

IG^® T7 RNA Polymerase functional testing was done by performing in vitro transcription with the IG^® sgRNA synthesis kits (Figure 3, also see Intact Genomics Catalog numbers 3203 and 3303.)

Figure 3: Synthesized RNA product analyzed on a Urea/TBE-PAGE RNA gel.

Lane 1: RNA ladder

Lane 2: IG synthesized guide RNA (60 bases)

Technical Support

Intact Genomics is committed to supporting the worldwide scientific research community by supplying the highest quality reagents. Each new lot of our products is tested to ensure they meet the quality standards and specifications designated for the product. Please follow the instructions carefully and contact us if additional assistance is needed. We appreciate your business and your feedback regarding the performance of our products in your applications.

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