Ribonucleic acid (RNA) condensation by thermal cycling with metal cations: yield of nanoparticles and their applicability for transfection

Abstract

To the present, different efficient but expensive, multistage, and time-consuming technologies have been developed to deliver ribonucleic acids (RNA) into eukaryotic cells. Here, we report a simple and feasible solution to design RNA nanocarriers based on nucleic acid condensation by bi- and trivalent metal ions during thermal cycling. Efficient RNA conversion to nanoparticles with small size (10–50 nm) suitable for transfection was achieved using cations Ni2+, Co2+ or Cu2+ alone or in combination with Ca2+ at the specially selected concentrations (2.0 mM–3.5 mM), low ionic strength, and narrow pH range (8.0–8.5). Other ions – Mn2+, Zn2+, Tb3+, or Gd3+ – caused RNA-cleaving effect that was abolished in the presence of Ni2+, Co2+, Zn2+, or Cu2+. Naked RNA-metal ion nanoparticles were extremely unstable in phosphate buffer and sensitive to serum ribonucleases (RNases), and this problem was solved by treatment with polyarginines-16 and 8. Polyarginine-stabilized nanoparticles, containing malachite green (MG) aptamer RNA and metal cations, crossed the cell membrane, dissociated in the cytoplasm, and preserved the functionality of transported RNA, as judged from efficient transfection of human embryonic kidney 293 cells. The technology, involving RNA condensation by metal cations, can be used as a cheap alternative to produce nanoscale carriers to deliver various RNAs into cells in vitro and in vivo.Communicated by Ramaswamy H. Sarma