Template Vector (BspQ I) for T7 mRNA Synthesis
This prelinearized vector contains essential elements for in vitro transcription (IVT), including a T7 promoter, transcription start sequence (AGG), 5′-untranslated region (UTR), 3′-UTR, poly(A) sequence, and BspQ I (Cat. # 1227) digestion site, all designed for high IVT performance.
This pre-linearized vector contains essential elements for in vitro transcription (IVT), including a T7 promoter, transcription start sequence (AGG), 5′-untranslated region (UTR), 3′-UTR, poly(A) sequence, and BspQ I (Cat. # 1227) digestion site, all designed for optimal high IVT performance (see Overview, Figure 1).
What sets this vector apart is its compatibility with a type IIS restriction enzyme BspQ I, which cleaves DNA sequences outside its recognition site (Overview, Figure 2). Using BspQ I for plasmid DNA linearized prior to in vitro transcription preserves the integrity of the poly(A) sequence located at the 3′ end of the IVT template. This is achieved by preventing the retention of additional base pairs from the restriction site. The intact poly(A) sequence is essential to achieve highly efficient translation in various mammalian cells, including human and murine cells.
Overview
Figure 1. Overview of Template Vector (BspQ I) for T7 mRNA Synthesis. This prelinearized vector contains essential elements for IVT reaction, including a T7 promoter (T7 p.), transcription start sequence (AGG), 5’-untranslated region (5’-UTR), 3’-UTR, poly(A) sequence, and BspQ I digestion site (red). To construct an IVT template plasmid, simply assemble a desired target coding sequence (Target CDS) using In-Fusion Cloning sites (green).
Figure 2. BspQ I restriction enzyme cleaves the Template Vector (BspQ I) for T7 mRNA Synthesis outside the recognition site. This preserves the integrity of the poly(A) tail sequence, which facilitates successful translation downstream of the in vitro transcription.
More Information
Applications
This plasmid vector has been designed for seamless assembly of your target coding sequence for in vitro transcription using In-Fusion Snap Assembly (Cat. # 638947, sold separately). The transcription start sequence (AGG) in this vector enables it to be coupled with cap analogs such as CleanCap Reagent AG or CleanCap Reagent AG (3′ OMe) from TriLink BioTechnologies.
For detailed information on the design of the template plasmid and procedures, please refer to the user manual of Cloning Kit for mRNA Template (Cat. # 6143) or visit our mRNA synthesis learning center.
Once the template plasmid is linearized using BspQ I, we strongly recommend utilizing the Takara IVTpro T7 mRNA Synthesis Kit (Cat. # 6144) as a high-yield system for in vitro transcription.
Both the Cloning Kit for mRNA Template and Takara IVTpro T7 mRNA Synthesis Kit are sold together as part of the Takara IVTpro mRNA Synthesis System (Cat. # 6141).
Additional product information
Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Certificates of Analysis and vector sequence information are located under the Documents tab.
In-Fusion cloning
Take advantage of simple, seamless technology that provides cloning efficiency over 95%, so that you get the right clone every time.
BspQ I
BspQ I creates linearized, scarless in-vitro transcription templates from plasmid DNA.
Takara IVTpro mRNA Synthesis System
A complete solution for synthesizing high-quality single-stranded mRNA containing your gene of interest with a cap structure and poly(A) sequence.
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