EZ Cap™ Cy5 Firefly Luciferase mRNA (5-moUTP)

The 5′ Cap structure protects mRNA from degradation and is required for ribosome recognition and translation initiation.

Cap 1, as a next-generation cap analog, closely mimics the natural structure of endogenous eukaryotic mRNA and is therefore recognized by cells as “self RNA.” This effectively reduces innate immune activation and enables efficient protein expression.

All APExBIO EZ Cap™ mRNA products utilize Cap 1 technology.

In addition to 5’ Cap, a complete in vitro transcribed (IVT) mRNA includes:

• • Optimized 5′ and 3′ untranslated regions (UTRs) to enhance stability and translation.
• • A Poly(A) tail (typically ~100 nucleotides) protects mRNA and facilitates translation.
• • Suitable nucleotide modifications based on the downstream applications.
• • Codon optimization based on the host expression system.

• m1Ψ (N1-methyl-pseudouridine): Strongly reduces innate immune responses and enhances protein expression. Widely used in mRNA vaccines research.
• ψUTP (pseudouridine), 5-moUTP (5-methoxyuridine), and 5mCTP (5-methylcytidine): Reduce immunogenicity while improving mRNA stability and translational efficiency. 5mCTP is commonly used in saRNA.
• Fluorescent- or Biotin-labeled Nucleotides (e.g. Cy5-UTP, Biotin-UTP): Used for visualization of mRNA to track mRNA delivery and localization.

RNA is highly susceptible to degradation.

• Thawing: Before use, allow the product to thaw completely on ice.
• Aliquoting: Briefly centrifuge and aliquot upon first use to avoid repeated freeze–thaw cycles.
• RNase Contamination Control: Use RNase-free consumables and reagents; maintain a clean and RNase-free working environment.
• Transfection: Always mix mRNA with transfection reagent first before adding to serum-containing medium to prevent RNase degradation.

• Check RNA Integrity: Make sure the RNA has been properly stored and handled according to the instruction, and avoid repeated freeze–thaw cycles.
• Check Transfection Procedure: Use reporter mRNA (such as EZ Cap™ Firefly Luciferase mRNA) as a positive control to evaluate transfection reagents, cell conditions and lab protocols.
• Optimize Experiments: Consider adjusting the RNA-to-transfection reagent ratio, cell density, incubation time, and protein detection time points etc.
• If the issue can’t be resolved, please contact our technical support team with detailed data for further assistance.

All RNA products are shipped on dry ice to maintain product integrity. If the dry ice has melted (and the product feels warm) or if the package is damaged upon receipt, please take photos immediately and contact us as soon as possible.