Transfected plasmid DNA is incorporated into the nucleus via nuclear envelope reformation at telophase

Abstract

DNA transfection is an important technology in the life sciences, wherein nuclear entry of DNA is necessary to achieve expression of the exogenous DNA. Non-viral vectors and their transfection reagents are useful as safe tools for transfection. However, they have no effects for transfection of non-proliferating cells, the reason for which remains unknown. This study aimed to elucidate the mechanism by which transfected DNA enters the nucleus for gene expression. To monitor the intracellular behavior of transfected DNA, we introduced a plasmid bearing lacO repeats and RFP-coding sequences into cells expressing GFP-LacI and observed plasmid behavior and RFP expression. RFP expression appeared only after mitosis. Electron microscopy showed that plasmids were wrapped with nuclear envelope (NE)-like membranes or associated with chromosomes at telophase. Depletion of BAF, which is involved in NE reformation, delayed plasmid RFP expression. These results suggest that transfected DNA is incorporated into the nucleus via NE reformation at telophase.

Enhanced Gene Delivery Triggered by Dual pH/Redox Responsive Host‐Guest Dimerization of Cyclooligosaccharide Star Polycations

“…In the case of SS‐bisAd , however, the high concentration of glutathione (GSH) in the cytosol would promote disulfide bridge cleavage, thereby deactivating the vector for strong pDNA recapture. This is critical for enabling subsequent nuclear entry of pDNA: although some examples have demonstrated the presence of cationic polymer molecules in the nucleus during polyplex‐mediated transfection, [ 21 ] the current evidence supports that pDNA must be essentially free or loosely bound to undergo either integration into the nucleus during nuclear envelop reformation at telophase in dividing cells [ 22 ] or, eventually, active import through the nuclear pore complex [ 23 ] ( Figure ). In CT2 derivatives the cavity is collapsed; [ 24 ] dissimilarities in pDNA complexation and transfecting properties between 1 and 2 in combination with the ditopic adamantane derivatives should then be principally ascribed to the host‐guest component of the process.…”

3D Confinement-enabled Priming of Synaptic Activation Promotes Primary T Cell Expansion

“…This expansion during cell division triggers physical enlargement, causing protrusions to extend from the cellular membranes and from the nucleus. The significant elevation of GFP expression following transfection suggests that 3D CPS could contribute to the rapidly growing field of CRISPR-based clinical trials, including those examining targeted knockout of immunosuppressive factors such as PD-1, of endogenous TCR and MHC-1, or studies examining CAR insertion among autologous and allogeneic T cells (47)(48)(49).…”

Lipofection with Lipofectamine™ 2000 in a heparin‐free growth medium results in high transfection efficiency in chicken primordial germ cells

“…Plasmid DNA is absorbed into the nucleus during cell division, and the transgene is expressed. Plasmid DNA transfected into the cytoplasm is incorporated into the nucleus during nuclear envelope reformation at the telophase [32] . Cell cycle synchronization and nuclear membrane destabilization facilitate plasmid DNA transfer to the nuclear membrane, thus improving transfection efficiency [33] .…”