Abstract:
:Small interfering RNAs (siRNAs) are now established as the preferred tool to inhibit gene function in mammalian cells yet trigger unintended gene silencing due to their inherent miRNA-like behavior. Such off-target effects are primarily mediated by the sequence-specific interaction between the siRNA seed regions (position 2-8 of either siRNA strand counting from the 5'-end) and complementary sequences in the 3'UTR of (off-) targets. It was previously shown that chemical modification of siRNAs can reduce off-targeting but only very few modifications have been tested leaving more to be identified. Here we developed a luciferase reporter-based assay suitable to monitor siRNA off-targeting in a high throughput manner using stable cell lines. We investigated the impact of chemically modifying single nucleotide positions within the siRNA seed on siRNA function and off-targeting using 10 different types of chemical modifications, three different target sequences and three siRNA concentrations. We found several differently modified siRNAs to exercise reduced off-targeting yet incorporation of the strongly destabilizing unlocked nucleic acid (UNA) modification into position 7 of the siRNA most potently reduced off-targeting for all tested sequences. Notably, such position-specific destabilization of siRNA-target interactions did not significantly reduce siRNA potency and is therefore well suited for future siRNA designs especially for applications in vivo where siRNA concentrations, expectedly, will be low.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Bramsen JB,Pakula MM,Hansen TB,Bus C,Langkjær N,Odadzic D,Smicius R,Wengel SL,Chattopadhyaya J,Engels JW,Herdewijn P,Wengel J,Kjems Jdoi
10.1093/nar/gkq341subject
Has Abstractpub_date
2010-09-01 00:00:00pages
5761-73issue
17eissn
0305-1048issn
1362-4962pii
gkq341journal_volume
38pub_type
杂志文章abstract::G-quadruplex (G4) DNA structures are linked to key biological processes and human diseases. Small molecules that target specific G4 DNA structures and signal their presence would therefore be of great value as chemical research tools with potential to further advance towards diagnostic and therapeutic developments. Ho...
journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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