Article ; Online: Design of Bifunctional Antisense Oligonucleotides for Exon Inclusion.
Methods in molecular biology (Clifton, N.J.)
2022 Volume 2434, Page(s) 53–62
Abstract: Bifunctional antisense oligonucleotide (AON) is a specially designed AON to regulate pre-messenger RNA (pre-mRNA) splicing of a target gene. It is composed of two domains. The antisense domain contains sequences complementary to the target gene. The tail ...
| Abstract | Bifunctional antisense oligonucleotide (AON) is a specially designed AON to regulate pre-messenger RNA (pre-mRNA) splicing of a target gene. It is composed of two domains. The antisense domain contains sequences complementary to the target gene. The tail domain includes RNA sequences that recruit RNA binding proteins which may act positively or negatively in pre-mRNA splicing. This approach can be designed as targeted oligonucleotide enhancers of splicing, named TOES, for exon inclusion; or as targeted oligonucleotide silencers of splicing, named TOSS, for exon skipping. Here, we provide detailed methods for the design of TOES for exon inclusion, using SMN2 exon 7 splicing as an example. A number of annealing sites and the tail sequences previously published are listed. We also present methodology of assessing the effects of TOES on exon inclusion in fibroblasts cultured from a SMA patient. The effects of TOES on SMN2 exon 7 splicing were validated at RNA level by PCR and quantitative real-time PCR, and at protein level by western blotting. |
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| MeSH term(s) | Exons/genetics ; Humans ; Oligonucleotides/chemistry ; Oligonucleotides, Antisense/genetics ; Oligonucleotides, Antisense/metabolism ; RNA Splicing ; RNA-Binding Proteins/metabolism |
| Chemical Substances | Oligonucleotides ; Oligonucleotides, Antisense ; RNA-Binding Proteins |
| Language | English |
| Publishing date | 2022-02-25 |
| Publishing country | United States |
| Document type | Journal Article ; Research Support, Non-U.S. Gov't |
| ISSN | 1940-6029 |
| ISSN (online) | 1940-6029 |
| DOI | 10.1007/978-1-0716-2010-6_3 |
| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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