Temperature controlled RNA isolation by N-isopropylacrylamide–vinylphenyl boronic acid copolymer latex

Abstract.

Thermosensitive N-isopropylacrylamide-4-vinylphenylboronic acid (NIPA-co-VPBA) copolymer latex particles were used as pseudospecific sorbent for the selective isolation of RNA by boronate affinity chromatography. In the proposed isolation procedure, RNA was adsorbed onto the latex particles via the interaction between boronic acid groups of the particles and diol groups of RNA at a low temperature (i.e. +4 °C). No significant amount of DNA was bound to the particles under the conditions that were used for RNA adsorption. This result indicated that the developed sorbent could be used effectively for selective removal of RNA from the RNA–DNA mixtures. RNA adsorption onto the latex particles significantly decreased with increasing temperature. On the other hand, NIPA-co-VPBA copolymer latex showed a thermoflocculation behaviour at temperatures higher than 30 °C. These two properties were used for temperature controlled RNA isolation by the proposed sorbent. After adsorption of RNA onto the stable latex particles at +4 °C, the particles were transferred into a desorption medium. Temperature increase in this medium resulted in both desorption of RNA from the particles and thermoflocculation of the latex suspension. Hence RNA was recovered in the clear supernatant without applying any additional separation for the removal of sorbent material. In the proposed procedure, the temperature was used as an on–off switch controlling the adsorption and desorption of RNA. The necessity of a separate medium (at a certain pH and ionic strength) for desorption was eliminated by the proposed isolation method. This behaviour allowed the desorption of RNA to a medium at any ionic strength and at any pH.