Related Experiment Video
Updated: Aug 23, 2026

Evaluation of Exon Inclusion Induced by Splice Switching Antisense Oligonucleotides in SMA Patient Fibroblasts
Published on: May 11, 2018
In vivo selection reveals combinatorial controls that define a critical exon in the spinal muscular atrophy genes
Natalia N Singh1, Elliot J Androphy, Ravindra N Singh
1Department of Medicine (LRB 326), University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-2324, USA. Ravindra.Singh@umassmed.edu
Abstract:
Humans have two near identical copies of the survival of motor neuron (SMN) gene, SMN1 and SMN2. In spinal muscular atrophy (SMA), SMN2 is not able to compensate for the loss of SMN1 due to an inhibitory mutation at position 6 (C6U mutation in transcript) of exon 7. We have recently shown that C6U creates an extended inhibitory context (Exinct) that causes skipping of exon 7 in SMN2. Previous studies have shown that an exonic splicing enhancer associated with Tra2 (Tra2-ESE) is required for exon 7 inclusion in both SMN1 and SMN2. Here we describe the method of in vivo selection that determined the position-specific role of wild-type nucleotides within the entire exon 7. Our results confirmed the existence of Exinct and revealed the presence of an additional inhibitory tract (3'-Cluster) near the 3'-end of exon 7. We also demonstrate that a single nucleotide substitution at the last position of exon 7 improves the 5' splice site (ss) such that the presence of inhibitory elements (Exinct as well as the 3'-Cluster) and the absence of Tra2-ESE no longer determined exon 7 usage. Our results suggest that the evolutionary conserved weak 5' ss may serve as a mechanism to regulate exon 7 splicing under different physiological contexts. This is the first report in which a functional selection method has been applied to analyze the entire exon. This method offers unparallel advantage for determining the relative strength of splice sites, as well as for identifying the novel exonic cis-elements.
Related Concept Videos
Exon Recombination
Exon shuffling follows “splice frame rules.” Each exon has three reading...
In-vitro Mutagenesis
Alternative RNA Splicing
There are five types of alternative RNA splicing that vary in the ways the pre-mRNA segments are removed or retained in the mature mRNA. The first...
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
RNA Splicing
Nucleotide Excision Repair
