Knockdown of p53 by Accell Self-delivering siRNA Causes Inhibition of p53-dependent DNA Damage Response in IMR-32 Neuroblastoma Cell Line and β-amyloid Toxicity in Rat Cortical Neurons
Poster Jun 12, 2015
Žaklina Strezoska, Tamara Seredenina1, Devin Leake, Annaleen Vermeulen
Neuroblastoma cell lines and primary neuronal cultures are commonly used as cellular model systems for studying cancer and neuronal development as well as being highly relevant models for the study of neurodegenerative diseases. However, most neuroblastoma cell lines and practically all primary neuronal cells suffer from low transfection efficiency due to the refractory nature of the cells to lipid-based transfection reagents. As such, application of siRNA for inducing RNA interference (RNAi), has limited utility in these cell types; thus limiting their usefulness for development of functional assays for screening and discovery of novel disease-relevant genes.
Dharmacon™ Accell™ siRNA enables efficient delivery in a wide range of cell lines and primary cells. Accell siRNA reagents carry a novel chemical modication pattern that facilitates the delivery of siRNA without a need for transfection reagents. To demonstrate the utility of Accell siRNA reagents in neuronal cells, the effects of the down-regulation of the tumor suppressor p53 was examined. This gene plays a pivotal role in mediating DNA damage-induced apoptosis as well as conferring a protective effect from β-amyloid peptide-induced neurotoxicity. Here we describe how application of Accell siRNA enabled the development of a high content screening assay in IMR-32 neuroblastoma cells and a whole culture cell viability assay in primary rat cortical neurons. The ability to modulate gene expression in neuronal cell lines and primary neurons using Accell siRNA opens new opportunities for functional genomic siRNA screens in the eld of neuroscience.
Multiplexing cell-based assays is possible using 3D culture models that are larger and more complex than monolayers
Real-time detection methods to measure live or dead cells provide much flexibility for multiplexing
All multiplexed assay combinations should be verified using appropriate controls for each 3D cell culture model.