Nucleoporation of mRNA

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Nucleoporation of mRNA

Nucleoporation has become an attractive alternative for the transfection of mRNA into primary and stem cells because it does not require the design of viral structures but directly deliver cargos to the cell nucleus with high gene transfer efficiency. Recently, this technique has been successfully applied for disease research and therapeutic development, including the research for the pathogenesis of the disease, gene therapy, immunotherapy, and stem cell generation. Creative Biolabs is an outstanding service provider who offers the most comprehensive and professional mRNA research strategies to advance clients' projects.

Introduction of Nucleoporation

Nucleoporation, also known as nucleofection, is used as a nonviral transfection method that delivers nucleic acids straight into the nucleus. This technique is a new electrotransfer technology that combines specialized solutions and electrical pulses to enable nucleic acid directly to transfer into the cell nucleus by electroporation. It is mainly used to transfect primary cell types which divide very slowly or do not divide at all. Primary cells are generally difficult to transfect by common methods that are strictly dependent on cell division, such as lipofection, while nucleoporation is suitable for primary cells since it is independent of cell division.

Introduction of Nucleoporation.

Hamm et al. utilized Nucleofector™ technology to transfect primary human melanocytes, human coronary smooth muscle cells, human chondrocytes, and human mesenchymal stem cells, and high efficiencies (28.9-45.3%) were observed. A plasmid expressing green fluorescent protein (GFP) was nucleoporated into monocyte-derived dendritic cells (DCs) using a high-throughput nucleofection system and transfection, and the efficiency was over 50%. These data suggest that nucleoporation is a promising non-viral transfection system for nuclear transfection of primary cells, providing a novel opportunity for gene therapy and stem cell research.

Flow cytometric detection of transgene expression 24h after nucleofection and conventional electroporation. Fig.2 Flow cytometric detection of transgene expression 24h after nucleofection and conventional electroporation. (Siemen, 2005).

Applications of Nucleoporation

Currently, nucleoporation is widely used for primary cells because of its unique features. This technique has been successfully used to transfect primary cells, cell lines, and stem cells. Primary human melanocytes, human coronary smooth muscle cells, human chondrocytes, and human mesenchymal stem cells were both non-virally transfected with higher efficiency compared to other non-viral methods, and the viability of the transfected cells was between 11.2% to 75%. Interestingly, nucleoporation can be used for genetic manipulation of DCs, which provides a new way in the development of vaccines for the treatment of cancer, chronic, and infectious diseases. Therapeutic or immunogenic genetic material could transfer into monocyte-derived immature and mature DCs with efficiencies approaching 60%. However, it is noted that the use of nucleoporation may result in the gradual loss of cell viability.

Advantages

  • Independent of cell division, suitable for hard-to-transfect cells such as suspension cells, primary cells, stem cells, and cell lines
  • A non-viral transfection method, without the design of viral constructs
  • With high transfection efficiency, the exogenous genes are directly transferred into the nucleus
  • An easy, fast, and efficient method
  • Maintain the physiological function of cells after transfection

Based on the nucleoporation technical platform, we provide the best and affordable services for mRNA transfection of primary cells, stem cells, and cell lines, with a high transfection efficiency and preferable cell viability. Our services will facilitate many different lines of research, including functional and structural genomics, drug discovery, and gene therapy. Please feel free to contact us and discuss your project.

Reference

  1. Siemen, H.; et al. Nucleofection of human embryonic stem cells. Stem Cells & Development. 2005, 14(4):378-83.
  2. Hamm, A.; et al. Efficient transfection method for primary cells. Tissue Engineering Part A. 2002, 8(2):235-245.
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