Antisense oligonucleotides, or ASOs, are 15–25 nt DNA sequences designed to bind complementary RNA targets, ultimately facilitating their degradation. ASO technology provided the first oligonucleotide-based approach to disrupting gene expression and has been used in knockdown experiments, target validation, drug therapy, and other applications. More recently, ASOs are used to study the role of long noncoding RNAs (lncRNAs) in gene regulation.
Many lncRNAs are localized to the nucleus. Often, lncRNA loss-of-function studies use techniques, such as RNAi, that are less effective because the nucleus contains a low amount of the required enzymes. ASOs, however, are particularly useful for studying nuclear lncRNAs, because ASOs engage RNase H—an enzyme prevalent in the nucleus that binds and cleaves DNA/RNA heteroduplexes.
IDT scientists have done extensive research on this application. See the Related reading list below for publications describing their findings.
RNase H1 is an endogenous nuclease that binds DNA/RNA heteroduplexes and cleaves the RNA strand, leading to degradation. For its action, cellular RNase H1 requires the heteroduplex to contain a segment of unmodified (with either native or phosphorothiate linkages) DNA that is ≥6 bases.
Modifications providing in vitro and in vivo stability
We recommend modifying your ASO sequence for increased stability and binding affinity. The most widely used modifications are phosphorothioate (PS) bonds, which are added throughout the oligonucleotide backbone to provide nuclease resistance. PS bonds have the potential to increase toxicity, which can be minimized by including an additional 2′-O-Methyl (2′OMe) RNA to create a “gapmer” ASO. DNA with these modifications display more nuclease resistance, lower toxicity, and increased hybridization affinities within cells and in vivo.
Ordering modified ASOs
To have phosphorothioate bonds added to your sequence, use an asterisk “*” between the bases.
2′-O-Methyl bases are represented with a lower case “m” in front of each base.
These oligos can be ordered with standard desalt or HPLC purification with Na+ salt exchange. The Na+ salt exchange is necessary to remove toxic salts from HPLC purification.
Click here to order IDT Antisense Oligonucleotides.