Morpholino Antisense Oligonucleotides: Understanding the Side Effects

Abstract

When a drug is advertised on television, it is always followed by a long list of possible side effects. Side effects are a familiar part of drug development and discovery, but they are also relevant to the field of biochemistry, particularly in regards to manipulation of gene expression. When a gene is knocked down, it is important that it is only the gene of interest that is being affected and not other non-target genes. When non-target genes are affected, the effect is referred to as ñoff-targetî. This off-target effect is a side effect of using morpholino antisense oligonucleotides, a commonly used reagent for knocking down genes in Danio rerio embryos. These off-target effects are believed to be caused by interactions with p53. However, based on previous data on patterns of expression for apex and its related proteins, the effects may be a result of excess oxidative damage to DNA. Robu et al. examined both on-target and off-target morpholinos for the developmental gene wnt5. The phenotypes resulting from both of these morpholinos have already been replicated in the Strauss Lab. RNA has been extracted, and the next step will be to conduct qRT-PCR to determine how the transcription of apex and its related genes vary between on-target and off-target injections. Protein levels will also be compared through western blotting. The results may show a deeper connection between off-target effects and oxidative damage to DNA with wide ranging implications for carcinogenesis and aging.