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In vitro Transcription

In vitro Transcription: INCOGNITO™ IVT and mRNA Production Systems

The new INCOGNITO™ kits

 

Evade cellular immune response  to your RNA with the new INCOGNITO™ kits  from Cellscript.

The new INCOGNITO™ kits *  are designed  for in vitro synthesis of RNA containing modified nucleosides such as pseudouridine and 5-methylcytidine. They assist in reducing immune response and enhancing protein translation.  A range of these kits are available to end users depending on their specific research desires and downstream applications.

Cellscript’s INCOGNITO™ contain everything needed to make N1meΨ- or Ψ-containing Cap1 mRNA, including in vitro transcription reagents plus ScriptCap™ Capping Enzyme, ScriptCap™ 2’-O-Methyltransferase, and A-Plus™ Poly(A) Polymerase.  

The INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit is specially formulated to enable users to obtain the maximum possible yields of N1-methyl-pseudouridine-containing RNA (N1meΨ-RNA) from an in vitro transcription (IVT) reaction. Although yield varies with the DNA template and other factors, a standard 60 minute, 20 μl reaction will yield up to 160 μg of RNA from 1 μg of the control template. These yields are made possible by the high-performance properties of the T7-FlashScribe™ enzyme. The standard reaction can be scaled up to produce milligram amounts of RNA containing the canonical nucleotides ATP, CTP, GTP and the modified nucleotide N1-methyl-pseudouridine-5'-triphosphate (N1meΨTP).

The INCOGNITO™ T7-FlashScribe™ Ψ-RNA Transcription Kit is specially formulated to maximize yields of pseudouridine-containing RNA (Ψ-RNA) from an in vitro transcription (IVT) reaction. Although yield varies with the DNA template and other factors, a standard 60 minute, 20 μl reaction will yield up to 160 μg of RNA from 1 μg of DNA template. These impressive yields are due to the high-performance properties of the T7-FlashScribe™ enzyme. The standard reaction can be scaled up to generate milligram amounts of RNA containing the canonical nucleotides ATP, CTP, GTP and the modified nucleotide pseudouridine-5'-triphosphate (ΨTP).

 

INCOGNITO™ T7 mScript™ mRNA Production Systems contain everything needed to make N1meΨ- or Ψ-containing Cap1 mRNA, including in vitro transcription reagents plus ScriptCap™ Capping Enzyme, ScriptCap™ 2’-O-Methyltransferase, and A-Plus™ Poly(A) Polymerase. 

 

The INCOGNITO™ SP6 Ψ-RNA Transcription Kit -optimized for high-yield synthesis of pseudouridine-containing RNA (Ψ-RNA) from an in vitro transcription (IVT) reaction.

The INCOGNITO™ T7 5mC- & Ψ-RNA Transcription Kit - optimized for high-yield synthesis of 5-methyl-cytidine- pseudouridine-containing RNA (5mCΨ-RNA) from an in vitro transcription (IVT) reaction.

Advantages

  • Optimized for high-yield synthesis of pseudouridine-containing IVT RNA (Ψ-RNA)
  • Produce up to 100 µg of Ψ-RNA from 1 µg of template ( depending on kit used).
  • Easily scale up reactions to produce milligram amounts of Ψ-RNA.
  • Lower immunogenicity in mammalian cells through incorporation of Ψ in synthesized mRNA.
  • Modified Ψ-mRNAs translate into higher protein levels
  • The standard reactions can be scaled up to produce milligram amounts of RNA canonical nucleotides ATP and GTP and modified pseudouridine-5'-triphosphate (ΨTP) and 5-methyl-cytidine-5'-triphosphate (5mCTP).


 

*Transcription Kits are for in vitro transcription of N1meΨ- or Ψ-containing RNA. - All kits above come with a label license to use them for internal research use under the University of Pennsylvania’s modified mRNA technologies patents.

INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit

INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit

CellScript

Catalogue No.DescriptionPack SizePriceQty
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IFMY240625INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit25 rxns POA Quantity
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  • Incorporates N1-methyl-pseudouridine into IVT RNA (N1meΨ-RNA) for reduced immunogenicity in mammalian cells.
  • Rapid, high yield production of up to 160 µg of N1meΨ-RNA from 1 µg of template in just one hour.
  • Easily scale up reactions to produce milligram amounts of N1meΨ-RNA.

Product is for research use only (RUO)

Use and Label Licenses for Specific Products: Purchaser receives a limited non-exclusive, non-transferable right to use the Products purchased from CELLSCRIPT™ solely for its own internal laboratory research use.

Product Description:

The INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit is specially formulated to maximize yields of N1-methyl-pseudouridine-containing RNA (N1meΨ-RNA) from an in vitro transcription (IVT) reaction. Although yield varies with the DNA template and other factors, a standard one hour, 20 μl reaction will yield up to 160 μg of RNA from 1 μg of DNA template. These high yields are due to the superior performance properties of the T7-FlashScribe™ enzyme. The standard reaction can be scaled up to generate milligram amounts of RNA containing the canonical nucleotides ATP, CTP, GTP and the modified nucleotide N1-methyl-pseudouridine-5'-triphosphate (N1meΨTP).

Modified N1meΨ-mRNAs translate into higher protein levels and lower innate immune responses in human and other mammalian cells that express various RNA sensors as compared to unmodified mRNAs.

INCOGNITO™ T7 IVT N1meΨ-RNA can be further processed into low immunogenicity mRNA (5'-end capped and 3'-end poly[A] tailed) through the use of CELLSCRIPT™'s ScriptCap™ Cap 1 Capping System (contains both ScriptCap™ Capping Enzyme and 2'-O-Methyltransferase) and A-Plus™ Poly(A) Polymerase Tailing Kit (available separately).

For maximum reduction of immunogenicity, follow production of INCOGNITO™ N1meΨ-mRNA with dsRNA removal from your sample using the Min-Immune™ Gold dsRNA Removal Kit to produce virtually dsRNA-free, ultra-low immunogenicity mRNA that is suitable for downstream applications such as cell and gene therapy research and mRNA vaccine development.

Product Performance:

The standard 60 minute, 20 μl reaction was optimized for transcription using 1 µg of linearized DNA template. However, incubation times can be adjusted if desired. Table 1 summarizes transcription results with 1 µg of the control template DNA in a standard reaction with incubation times from 20-120 minutes. Results may vary depending on the template used.

 

Table 1. IVT RNA yields from INCOGNITO™ T7-FlashScribe™ N1meΨ-RNA Transcription Kit reactions incubated between 20-120 minutes. The standard protocol uses an incubation time of 60 minutes.

 

 

Incubation Time (minutes) 20 30 60 90 120
RNA Yield (µg) 78-80 100-109 120-175 120-172 121-199

 

Materials Supplied

Important Store at -20°C in a freezer without a defrost cycle. Do not store at -70°C.

  

T7 Control Template DNA: Is a linearized 4.1 kb plasmid that contains a T7 promoter followed by a phage lambda dsDNA insert that encodes a 1,375 base runoff transcript.  The Control Template DNA is provided at a concentration of 0.5 µg/µl in T10E1 Buffer (10 mM Tris-HCl, pH 7.5, 1 mM EDTA).

Materials Required, but not Supplied

Terms and Conditions: The Product listed is currently offered by CELLSCRIPT™ for research use only under the defined Terms and Conditions.

 

Troubleshooting guide:


FAQs: For answers to Frequently Asked Questions about our products, visit the FAQ library.

If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200

Manual

SDS

These are both links to the manufacturer's page

If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200

  1. Karikó, K. et al., (2008) Molecular Therapy 16, 1833.
  2. Anderson, B.R. et al., (2010) Nucl. Acids Res. 17, 5884.
  3. Karikó, K. et al., (2005) Immunity 23, 165.
  4. Karikó, K. and Weissman, D. (2007) Curr. Opin. Drug Discov. Devel. 10, 523.
  5. Andries, O. et al., (2015) J. Controlled Release 217, 337.
  6. Svitkin, Y. V. et al., (2017) Nucleic Acids Res. 45 (10), 6023.

If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200

Uses and Label Licenses for Specific Products

INCOGNITO™ kits or other products or services comprising N1-methyl-pseudouridine, pseudouridine, 5-methyl-cytidine and/or other modified nucleotides for use in making RNA or mRNA (the “Products”) that are covered by technologies and IP disclosed in U.S. Patent Nos. 8,278,036 and 9,750,824 and PCT Publ. Nos. WO 2007/024708 and WO/2011/071931 and U.S. and international divisional, continuation, or other patents and patent applications derived therefrom that are owned by the University of Pennsylvania and licensed to CELLSCRIPT (“Products”) are sold for research use only. By purchasing these Products from CELLSCRIPT or an authorized distributor of CELLSCRIPT, Purchaser receives a limited non-exclusive, non-transferable right to use the Products purchased from CELLSCRIPT solely for its own internal laboratory research use (the “Product Use”). Such Product Use expressly excludes any Commercial Use comprising [A] clinical or other use in humans, [B] veterinary, livestock, agricultural or commercial use in animals, and/or [C] manufacture, distribution, importation, exportation, or sale of any products and/or services made using Products, it being understood that [A]–[C] are separately and/or collectively defined as a “Commercial Use” herein.

By purchase of Products, Purchaser agrees not to make, import, use, transfer, offer for sale, or sell Products, components of Products, or derivatives of Products, including modified nucleoside-containing RNA or cells made by use of Products, or to provide services, information, or data obtained by the use thereof in exchange for money or other consideration. CELLSCRIPT provides no warranties (statutory or implied) concerning non-infringement of intellectual property rights of any other parties, and all such warranties are expressly disclaimed. Please contact CELLSCRIPT to inquire about a license for a Commercial Use.

The license remains in effect as long as the product is being used in accordance with the license terms. The license is terminated if the user breaches any of the license terms, such as using the product for commercial purposes without proper authorization.

 

Important

Store at -20°C in a freezer without a defrost cycle. Do not store at -70°C.

If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200

If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200