Uptake of spherical nucleic acid (SNA) in Ochromonas danica: A new potential biotechnological tool

The development of a nanomaterial-based platform for application as an in vivo sensor or genome-editing tool is of great importance to microalgal biotechnology. In this study, gold nanoparticles (AuNPs) were functionalized with DNA to form spherical nucleic acid (SNA) through a pH-assisted method. The fabricated SNA readily entered Ochromonas danica, a microalga species that is suitable to study the endocytosis mechanisms, after the cells were simply incubated with SNA solution for 5 h. The transmission electron microscopy (TEM) images demonstrated the internalization of SNA within both the cytoplasm and vacuole, indicating their intracellular presence and distribution. The uptake of SNA was significantly inhibited by the clathrin-mediated endocytosis inhibitor (chlorpromazine, CPZ) and caveolae-mediated endocytosis inhibitor (methyl-beta-cyclodextrin, M beta CD), suggesting that SNA was taken up by O. danica cells mainly through the clathrin- and caveolae-dependent pathways. The toxic effects of SNA on O. danica were negligible since the growth and photosynthesis of treated cells were comparable to those of untreated cells (control). As a proof-of-concept, the SNA platform was used as an intracellular sensor for microalgae by using an aptamer-based SNA (i.e., aptamer nano-flare). The aptamer nano-flare fluorescently imaged the intracellular ATP, showing the high potential for SNA applications. This study will contribute to the application of nanotechnology to improve microalgal biotechnology.