DNA interpolyelectrolyte complexes as a tool for efficient cell transformation

Abstract

A tool was developed for enhancement of plasmid penetration into an intact cell, based on increasing DNA hydrophobicity via inclusion into a soluble interpolyelectrolyte complex (IPC) with polycations. The characteristics of formation of DNA IPC with synthetic polycations [poly(N-ethyl-4-vinylpyridinium)bromide (PVP) and PVP modified with 3% of N-cetyl-4-vinylpyridinium units (PVP-C)] were studied using ultracentrifugation and polyacrylamide gel electrophoresis methods. The conditions were established under which the mixing of DNA and polycation aqueous solutions results in the self-assembly of soluble IPC species. Incorporation of DNA into IPC results in the enhancement of DNA binding with isolated Bacillus subtilis membranes. A considerable increase in the efficiency of transformation of B. subtilis cells with pBC16 plasmid resulted from incorporation of the plasmid into the IPC with PVP and CVP.

DNA affinity to biological membranes is enhanced due to complexation with hydrophobized polycation

“…At the same time, it was shown in the experiments with living cells that complexation of plasmid DNA with both P2 and P2,16 considerably enhanced DNA affinity to living cell membranes [14][15][16]. Such a difference in the behavior of DNA-containing complexes regarding liposomes and cells probably resulted from the rigidity of small liposomes, of which the membranes could be hardly curved without disruption.…”

“…A number of approaches have been developed to introduce DNA into cells [2][3][4][5][6][7][8][9][10][11][12][13]. Recently, for this purpose, it was proposed to incorporate nucleic acids into soluble IPECs with polycations [11][12][13][14][15][16]. Incorporation of DNA into soluble IPEC with quaternized poly-4-vinylpyridine was shown to enhance significantly the efficiency of DNA penetration into Bacillus subtilis cells when compared to native DNA [14][15][16].…”

“…Recently, for this purpose, it was proposed to incorporate nucleic acids into soluble IPECs with polycations [11][12][13][14][15][16]. Incorporation of DNA into soluble IPEC with quaternized poly-4-vinylpyridine was shown to enhance significantly the efficiency of DNA penetration into Bacillus subtilis cells when compared to native DNA [14][15][16]. At the same time, it was shown that amphiphilic oligocations (such as spermine modified by hydrophobic substituents) complexed with DNA effectively stimulated gene transfer into mammalian cells [13].…”

Polymer-based gene delivery with low cytotoxicity by a unique balance of side-chain termini

“…In general, discrete polycation-polyanion complexes are formed spontaneously in aqueous solutions by ionic bonds between the polymers. Cooperativity of the ionic bonds depends on the density of the charge repeat units on each polymer with a minimum of 6 bonds, also called the lower critical chain length, required for a cooperative system (39)(40)(41). Given the randomized amidation of the imidazole on the polylysine in polymers 1-3, it is unlikely that the repeat cationic units are spaced equally throughout the polymer, potentially reducing the number of long cationic repeat units among the polymer side chains, thus reducing cooperativity between the polymer and DNA.…”

Design of amphiphilic oligopeptides as models for fine tuning peptide assembly with plasmid DNA

“…This is evident from the precipitation seen in the peptide-DNA complexes. In the stoichiometry complex, the negative charge of the DNA is completely compensated (14,15). As the concentration of the peptide further increases, the portion of the stoichiometry complex grows, while the non-stoichiometric complex falls down.…”