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Applications | mRNA and Viral RNA Transfection

High Efficiency Low Toxicity Large RNA Transfection

While smaller RNA species such as siRNA/miRNA, etc. are routinely transfected by researchers investigating gene function, transfection of large RNA molecules such as mRNA transcripts and viral RNA replicons is an emerging trend. mRNA delivery for protein expression offers an inherent advantage as cytoplasmic delivery is sufficient to initiate translation of the transcript. mRNA is a single stranded RNA molecule that encodes a protein and may have post-transcriptional modifications such as a 5’ cap and 3’ poly(A) tail. It can occur naturally in a mammalian cell or be produced in vitro to mimic natural mRNA. An in vitro transcript is an unmodified RNA molecule produced from a DNA template using one of the three phage DNA-dependent RNA polymerases (T7, T3, or SP6).

There have been tremendous improvement in in vitro transcription technology that facilitate transfection of RNA transcripts for prolonged and sustained protein expression. The TransIT®-mRNA Transfection Kit is formulated specifically for delivery of larger RNA species into mammalian cells. It consists of the TransIT®-mRNA Reagent-a novel non-liposomal cationic polymer/lipid formulation and the mRNA Boost Reagent- a proprietary compound. Both components are required to complex with RNA for efficient delivery into the cytoplasm. Salient features of this kit include:

  • High Efficiency RNA Delivery - Achieve RNA delivery in a large population of cells to ensure experimental success
  • Low Cellular Toxicity - Maintain cell density and reduce experimental biases
  • Serum Compatibility - Perform transfections in the presence of serum which eliminates the need for a media change and maintains cellular health
  • Delivery of Various Sizes of RNA - Ideal for specialized applications, such as stem cell reprogramming and RNA virus production
 

TransIT-mRNA Transfection Kit Mock Transfection and lacZ Data

 

TransIT®-mRNA Transfection Kit Efficiently Delivers lacZ mRNA to CHO-K1 Cells. Using the TransIT®-mRNA Transfection Kit, CHO-K1 cells were mock transfected (A) or transfected with a capped and polyadenylated lacZ encoding mRNA (B). Approximately 18 hrs post-transfection the cells were stained using Mirus Bio’s Beta-gal Staining Kit to identify the lacZ transfected cells.

 

TransIT-mRNA Transfection Kit Multiple Dendritic Cell Types Data

 

Multiple Dendritic Cell Types Express GFP from mRNA Transfected by TransIT®-mRNA Transfection Kit. Murine primary bone marrow derived dendritic cells (BMDC) and murine dendritic cells types (JAWSII and DC 2.4) were transfected with 1 µg of capped and polyadenlyated mRNA encoding GFP using a TransIT®-mRNA Reagent: Boost: mRNA ratio of 1:1:1 (µl:µl:µg). Primary BMDCs, JAWSII and DC 2.4 were seeded (80,000 cell/well) overnight in 24-well plates. Cells were assayed via flow cytometry 8 hours post transfection. Error bars represent the standard deviation of at least 3 separate experiments. Data courtesy of Kyle Phua (Principal Investigator: Kam W. Leong), Duke University

 

 

TransIT-mRNA Transfection Kit Easy Transfection Protocol Schematic
 

Save Time and Resources over Electroporation

  • Superior Performance, Low Toxicity
  • Easier-to Use TransIT®-mRNA Kit Saves Valuable Time
  • No Electroporator needed - Saves Valuable Resources

 

TransIT-mRNA Transfection Kit 10x Better than Electroporation Data

10X Better Performance Using 2.5X Less RNA. Huh7 cells were transfected with 2 µg of a yellow fever virus-luciferase expressing RNA replicon (YFV-luc) using the TransIT®-mRNA Transfection Kit or electroporated in parallel with 5 µg of the YFV-luc RNA replicon using an ECM630 Electroporator (BTX®) under optimal conditions. Twenty-four hours later, the cells were harvested and assayed for luciferase activity and total protein content. Luciferase activity was normalized to total protein content to correct for differences in cell number between samples (1).

(1) Gonzalez, G. et al. (2007) Selection of an Optimal RNA Transfection Reagent and Comparison to Electroporation for the Delivery of viral RNA. J. Virological. Methods145:14-21.

 

TransIT-mRNA Transfection Kit TransIT mRNA vs Electroporation-Time Saving Protocol Comparison

 

Simple Protocol Saves Over 4 Hours. Using TransIT®-mRNA saves more than four hours in comparison to electroporation.

Comparison Between TransIT-mRNA and Electroporation Highlights Where You Can Save Time, Budget and Materials

 TransIT®-mRNA KitElectroporation
Cell Number~3.8 x 105
(per well of 6-well plate)
Scalable based on culture vessel surface area
~5 x 106
(Plated in 6-well plate after electroporation)
RNA Amount2.5 µg
(per well of 6-well plate)
Scalable based on culture vessel surface area
~5 µg
(per standard electroporation)
Serum CompatibilityCompatible
- Enhances cell viability
- Simplifies protocol
Incompatible
- Requires serum free conditions
- Requires media changes

Products for mRNA Transfection

TransIT-mRNA Transfection Sample

TransIT®-mRNA Transfection Kit

Each Kit is supplied with the TransIT®-mRNA Transfection Reagent and the mRNA Boost Reagent

  • Low Cellular Toxicity – Maintain cell density and reduce experimental biases
  • High Efficiency Delivery – Achieve RNA delivery in a large population of cells to ensure experimental success
  • Serum Compatible – Perform transfections in the presence of serum which eliminates the need for a media change and maintains cellular health
  • Deliver Various Sizes of RNA – Ideal for specialized applications, such as virus production, protein expression and CRISPR genome editing