Comparison of three magnetic bead surface functionalities for RNA extraction and detection

Abstract

Magnetic beads are convenient for extracting nucleic acid biomarkers from biological samples prior to molecular detection. These beads are available with a variety of surface functionalities designed to capture particular subsets of RNA. We hypothesized that bead surface functionality affects binding kinetics, processing simplicity, and compatibility with molecular detection strategies. In this report, three magnetic bead surface chemistries designed to bind nucleic acids, silica, oligo (dT), and a specific oligonucleotide sequence were evaluated. Commercially available silica-coated and oligo (dT) beads, as well as beads functionalized with oligonucleotides complementary to respiratory syncytial virus (RSV) nucleocapsid gene, respectively recovered ∼75, ∼71, and ∼7% target RSV mRNA after a 1 min of incubation time in a surrogate patient sample spiked with the target. RSV-specific beads required much longer incubation times to recover amounts of the target comparable to the other beads (∼77% at 180 min). As expected, silica-coated beads extracted total RNA, oligo (dT) beads selectively extracted total mRNA, and RSV-specific beads selectively extracted RSV N gene mRNA. The choice of bead functionality is generally dependent on the target detection strategy. The silica-coated beads are most suitable for applications that require nucleic acids other than mRNA, especially with detection strategies that are tolerant of a high concentration of nontarget background nucleic acids, such as RT-PCR. On the other hand, oligo (dT) beads are best-suited for mRNA targets, as they bind biomarkers rapidly, have relatively high recovery, and enable detection strategies to be performed directly on the bead surface. Sequence-specific beads may be best for applications that are not tolerant of a high concentration of nontarget nucleic acids that require short RNA sequences without poly(A) tails, such as microRNAs, or that perform RNA detection directly on the bead surface.

Keywords: RNA extraction; nucleic acid extraction; oligo (dT) magnetic particles; sequence-specific magnetic particles; silica-coated magnetic particles.