Nucleic Acid Co-transfection

cotransfection

Co-transfection or co-delivery of multiple nucleic acids - either similar or different in nature is a technique routinely employed by cell biologists to address increasingly complex experimental design and applications. Common examples of nucleic acid co-transfection are as follows:

  • Co-delivery of multiple plasmid DNAs for virus production
  • Co-transfection of plasmid DNA and siRNA to investigate gene function
  • Co-delivery of plasmid DNA and mRNA for gene insertion, etc.

Most transfection reagents are designed for efficient delivery of a single nucleic acid type such as plasmid DNA, siRNA, mRNA, etc. Therefore, co-transfecting multiple nucleic acids of the same kind such as multiple plasmid DNAs or siRNAs is relatively straightforward as long as the ratio of the amount of transfection reagent to total nucleic acid is maintained. Co-delivery of completely different nucleic acid types can be quite tricky depending on the combination.

Co-transfection of different nucleic acid types such as large plasmid DNA and smaller siRNA/miRNAs can be challenging primarily due to the size and charge difference between these nucleic acid molecules. The charge of the nucleic acid molecule impacts the charge ratio of the transfection complexes and their formation and uptake by the mammalian cell membrane. Due to this reason, transfection reagents that can deliver smaller nucleic acids such as siRNA/miRNA to the cytoplasm efficiently are not always successful in plasmid DNA delivery to the nucleus. Unique polymeric formulations such as the TransIT-X2® Dynamic Delivery System are very effective nucleic acid condensing agents. These polymers provide positive charged side groups that can interact efficiently with the negative backbone of DNA and siRNA to form tight complexes that have a net positive surface charge. Once formed and condensed the transfection complexes are taken up very efficiently via endocytosis and nucleic acid delivery to their respective functional locations: the nucleus (DNA) and cytoplasm (siRNA/miRNA). Learn more about how the TransIT-X2® Dynamic Delivery System enables co-transfection of plasmid DNA and siRNA here.

For transfection reagent recommendations to deliver various different combinations of nucleic acids, please refer to the table here. If a single transfection reagent is not suitable for a specific co-delivery application, then separate transfection complexes can be prepared using the optimal transfection reagent for each nucleic acid and added to the cells at the same time. As long as the individual transfection protocols are compatible, this method ensures uptake of all nucleic acids by the cell simultaneously.

 

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