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HiScribe® T7 ARCA mRNA Kit (with tailing)

Catalog # Concentration Size List Price Quantity Your Price
E2060S 20 reactions $602.00
$541.80
Catalog # Size List Price Your Price
E2060S 20 reactions $602.00
$541.80
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca
  • Generate up to 25 μg of capped and tailed mRNA per reaction
  • mRNA capping, DNA removal, mRNA tailing and purification complete in 2 hours. Competitive products have more pipetting steps and separate components.
  • Enables partial incorporation of modified bases N1-Methyl-Pseudouridine-5’-Triphosphate (NEB #N0431), 5-Methyl-Cytidine-5’-Triphosphate (NEB #N0432), Pseudouridine-5’-Triphosphate (NEB #N0433) and 5-Methoxy-Uridine-5’-Triphosphate (NEB #N0434)
  • Ultra high-quality components ensure mRNA integrity
  • ARCA-based capping in correct orientation ensures high translation efficiency
  • Template removal and mRNA purification reagents included
  • HiScribe kits contain twice the number of reactions as competitive products
  • Also available without tailing reagents (NEB #E2065)
  • Getting ready to scale up RNA synthesis? Download our new technical note “Scaling of High-Yield In vitro Transcription Reactions for Linear Increase of RNA Production” for a generalized set of recommendations for synthesizing high yields of RNA.
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    Avoiding RNase Contamination
Most eukaryotic mRNAs require a 7-methyl guanosine (m7G) cap structure at the 5´ end and a poly(A) tail at the 3´ end to be efficiently translated. The HiScribe T7 ARCA mRNA Kit (with tailing) is designed for quick production of ARCA capped and poly(A) tailed mRNA in vitro. Capped mRNAs are synthesized by co-transcriptional incorporation of Anti-Reverse Cap Analog (ARCA, NEB #S1411) using T7 RNA Polymerase. The transcription reaction can be set up easily by combining the ARCA/NTP mix, T7 RNA Polymerase Mix and a suitable DNA template. The kit also allows for partial incorporation of 5mCTP, Pseudo-UTP and other modified nucleotides into mRNA. After a brief DNase I treatment to remove the template DNA, capped mRNA is poly(A) tailed with Poly(A) Polymerase. mRNAs synthesized with the kit can be used for cell transfection, microinjection, in vitro translation and RNA vaccines.

ARCA is incorporated into mRNA exclusively in the correct orientation, generating capped mRNA that is more efficiently translated. Standard cap analogs can be incorporated in either direction resulting in only 50% of capped mRNA that is functional in protein translation.

Figure 1. Structure of Anti-Reverse Cap Analog (ARCA, NEB #S1411)

Methylation at the 3´ position of 7mG forces the cap structure to be attached to mRNA in the correct orientation.
Figure 2. Overview of mRNA synthesis work flow with the HiScribe T7 ARCA mRNA Kit (with tailing)Figure 2. Overview of mRNA synthesis work flow with the HiScribe T7 ARCA mRNA Kit (with tailing)

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  Poly(A) Polymerase Reaction Buffer B0276SVIAL -20 1 x 1.5 ml 10 X
  T7 RNA Polymerase Mix M0255AAVIAL -20 1 x 0.04 ml
  ARCA/NTP Mix N2053AVIAL -20 1 x 0.2 ml
  DNase I (RNase-free) M0303AAVIAL -20 1 x 0.04 ml 2,000 units/ml
  CLuc Control Template N0247AVIAL -20 1 x 20 µl 0.25 mg/ml
  LiCl Solution B2051AVIAL -20 1 x 1.4 ml
  E. coli Poly(A) Polymerase M0444AVIAL -20 1 x 0.1 ml
  Dithiothreitol (DTT) B1222AVIAL -20 1 x 0.5 ml 100 mM

Properties & Usage

Materials Required but not Supplied

  • DNA template
  • Thermocycler or 37°C incubator.
  • Nuclease-free water
  • Buffer- or water-saturated phenol:chloroform
  • Ethanol
  • 3 M Sodium acetate, pH 5.2
  • 5 M Ammonium acetate
  • Spin columns (see Monarch® RNA Cleanup Kits, NEB #T2040 or #T2050)
  • Gels, running buffers and gel box
  • Equipment for RNA analysis


Companion Products
Notes
  • All kit components should be stored at –20°C. The kit contains sufficient reagents for 20 reactions of 20 μl each. Each standard reaction yields up to 20 μg of capped mRNA from 1 μg control template. Up to 25 μg capped and tailed mRNA can be obtained after Poly(A) tailing and purification by LiCl precipitation.
Citations
Publications
  • Marc Herb, Alexander Gluschko, Katja Wiegmann, Alina Farid, Anne Wolf, Olaf Utermöhlen, Oleg Krut, Martin Krönke, Michael Schramm (2019). Science Signaling. 12(568), eaar5926..
  • Marc Herb, Alina Farid, Alexander Gluschko, Martin Krönke, Michael Schramm (2019). Highly efficient transfection of primary macrophages with in vitro transcribed mRNA J Vis Exp. e60143..
Quality Control Assay
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
Certificate of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety Data Sheets
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This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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