Transfection types, methods and strategies: a technical review

Abstract

Transfection is a modern and powerful method used to insert foreign nucleic acids into eukaryotic cells. The ability to modify host cells' genetic content enables the broad application of this process in studying normal cellular processes, disease molecular mechanism and gene therapeutic effect. In this review, we summarized and compared the findings from various reported literature on the characteristics, strengths, and limitations of various transfection methods, type of transfected nucleic acids, transfection controls and approaches to assess transfection efficiency. With the vast choices of approaches available, we hope that this review will help researchers, especially those new to the field, in their decision making over the transfection protocol or strategy appropriate for their experimental aims.

Keywords: Chemicals; Controls; Efficiency; Methods; Nucleic acids; Transfection.

Figure 1. The workflow of literature selection using PRISMA flow strategy.

Keywords used during the literature search included “transfection”, “co-transfection”, “chemicals”, “reagents”, “DNA”, “siRNA”, “shRNA”, “miRNA”, “plasmid”, “oligonucleotides”, “efficiency”, “safety”, “cytotoxicity” and “controls”. Inclusion criteria of the literature selection comprised of English written articles or sources which were reported in the past 30 years since 1990 and articles which reported the description or comparison between different transfection protocols, types of transfected nucleic acids, transfection controls, methods to assess transfection efficiencies and transfection reagents in terms of their strengths, effectiveness, safety level and/or limitations. About 150 written sources were used in the qualitative analysis of this technical review.

Figure 2. Different transfection protocols that can be divided into viral-based, non-viral based or combination of both (hybrid).

Figure 3. Summary of the commonly used chemical transfection reagents.

Transfection reagents can be generally divided into liposomal or high-lipid based and non-liposomal based reagents. Non-liposomal reagents can be mixed protein-lipid reagents, non-liposomal lipids or non-protein non-lipid reagents like dendrimer. The details of the Invitrogen and ThermoFisher Scientific products can be found at: https://www.thermofisher.com/my/en/home.html. Information on the Qiagen related transfection products can be found at: https://www.qiagen.com/us/. Information on the Promega related reagents can be found at: https://worldwide.promega.com/. For Polyplus related transfection reagents, the information can be retrieved at: https://www.polyplus-transfection.com/. For Sigma–Aldrich or Merck or Roche related reagents, the information can be found at: https://www.sigmaaldrich.com/united-states.html. Information on the Thomas Scientific related chemicals can be retrieved at: https://www.thomassci.com/. Details on the Mirus Bio products can be referred at: https://www.mirusbio.com/. For GeneCopoeia products, further information can be found at: https://www.genecopoeia.com/. Information on ClonTech products can be found at: https://www.takarabio.com/. Details on System Biosciences chemicals can be found at: http://www.excellbio.com/. Information on the products by Strategene can be found at: https://www.agilent.com/. Details of the Fermentas International Inc. can be found at: http://fermentas.lookchem.com/.

Figure 4. Applications of co-transfection in biotechnology and life sciences research.

(A) Multiple plasmids co-transfection is useful in generating a hybrid vector and is for protein–protein interaction studies. (B) Multiple small RNAs co-transfection is popular in RNA interference and functional assay study to evaluate the regulatory effects of the small RNA on the expression of the downstream target. (C) Co-transfecting a small RNA molecule and a plasmid DNA that carries a reporter system can be used to assess small RNA transfection efficiency or to determine the regulatory effects of the small RNA on a specific gene.