Transfection

For work with bacterial and archaeal cells transfection retains its original meaning as a special case of transformation.

[5] Physical methods include electroporation, microinjection, gene gun, impalefection, hydrostatic pressure, continuous infusion, and sonication.

Chemicals include methods such as lipofection, which is a lipid-mediated DNA-transfection process utilizing liposome vectors.

[5] Other physical methods use different means to poke holes in the cell membrane: Sonoporation uses high-intensity ultrasound (attributed mainly to the cavitation of gas bubbles interacting with nearby cell membranes), optical transfection uses a highly focused laser to form a ~1 μm diameter hole.

[9] Chemical-based transfection can be divided into several kinds: cyclodextrin,[10] polymers,[11] liposomes, or nanoparticles[12] (with or without chemical or viral functionalization.

Following this, fusogenic agents (e.g., Sendai virus, PEG, electroporation) are used in order to fuse the protoplast carrying the gene of interest with the target recipient cell.

As with DNA, RNA can be delivered to cells by a variety of means including microinjection, electroporation, and lipid-mediated transfection.

[31] Long-RNA transfection is the process of deliberately introducing RNA molecules longer than about 25nt into living cells.

A distinction is made between short- and long-RNA transfection because exogenous long RNA molecules elicit an innate immune response in cells that can cause a variety of nonspecific effects including translation block, cell-cycle arrest, and apoptosis.

The innate immune system has evolved to protect against infection by detecting pathogen-associated molecular patterns (PAMPs), and triggering a complex set of responses collectively known as inflammation.

The specific chemical, structural or other characteristics of long RNA molecules that are required for recognition by PRRs remain largely unknown despite intense study.

At any given time, a typical mammalian cell may contain several hundred thousand mRNA and other, regulatory long RNA molecules.

Although only a very small number of these modified nucleotides are present in a typical mRNA molecule, they may help prevent mRNA from activating the innate immune system by disrupting secondary structure that would resemble double-stranded RNA (dsRNA),[46][34] a type of RNA thought to be present in cells only during viral infection.

Inhibiting only three proteins, interferon-β, STAT2, and EIF2AK2 is sufficient to rescue human fibroblasts from the cell death caused by frequent transfection with long, protein-encoding RNA.

[45] Inhibiting interferon signaling disrupts the positive-feedback loop that normally hypersensitizes cells exposed to exogenous long RNA.