DNA/2'-O-Methyl RNA Chimera

Incorporating 2'-O-methylRNA bases into antisense oligonucleotides or miRNAs can enhance their stability, prevent them from being degraded by nucleases, and increase their affinity for target mRNA by increasing Tm. With years of experience in custom oligonucleotide modification, Creative Biolabs provides a comprehensive range of oligonucleotide modification services, including 2'-O-methyl ester modification. Our scientists are proficient in the preparation of DNA/2'-O-Methyl RNA chimera with high purity and high quality to promote clients' meaningful research.

Core Composition and Modification of DNA/2'-O-Methyl RNA Chimera

• DNA Unit
  composed of deoxyribonucleotides, with a deoxyribose backbone, consistent with the structure of natural DNA, and binds to target nucleic acids (such as mRNA and DNA) through complementary base pairing.
• 2'-O-MethylRNA Unit
  The 2'-hydroxyl group of RNA nucleotides is methylated (i.e. 2'-O-Me RNA). This modification does not alter the base pairing ability, but significantly enhances the stability of the molecule - resisting degradation by nucleases such as RNase and reducing its likelihood of being recognized by the human immune system as an "exogenous nucleic acid".
• Chimera
  The two are connected by a phosphodiester bond. A common design is the "DNA core region+2'-O-methylRNA end region" (e.g., modified RNA at both ends and DNA in the middle) to balance binding specificity and in vivo stability.

Introduction of 2'-O-Methyl RNA

2'-O-methylation is a common nucleoside modification of RNA, in which a methyl group is added to the 2' hydroxyl of the ribose moiety of a nucleoside to produce a methoxy group. 2'-O-Methyl RNA is a naturally occurring modification of RNA found in tRNA and small RNAs. It represents one of the earliest sugar modifications employed in antisense medicinal chemistry and is widely used to improve the nuclease stability of siRNA or antisense oligonucleotides. A number of studies have revealed that the oligonucleotides modified with 2'-O-Methyl RNA showed higher nuclease resistance and affinity to target mRNA compared to unmodified oligonucleotides. Especially, the combination of 2'-O-Methyl RNA with phosphorothioate linkages is recommended to be introduced in the process of oligos synthesis in order to further increase the nuclease resistance.

Figure 1 Model of 2′-O-methylation (2′-O-Me) profile modulation and its consequence on the intrinsic translational activity of ribosomes.¹

How to Create an RNA Chimera?

The synthesis of DNA/2'-O-methylRNA chimeras is a complex process based on mature solid-phase synthesis protocols. This process requires highly skilled chemists and specialized equipment.

The basic steps are as follows:

1. Solid Support Attachment: covalently attach the first nucleotide to a solid phase carrier, such as controlled pore size glass (CPG).
2. Deprotection: Deprotech the 5 '- hydroxyl group of the first nucleotide for the next step of conjugation.
3. Coupling: Activate the phosphoramide structural unit (DNA or 2'-O-methylRNA phosphoramide) and couple it with the deprotected 5'-hydroxyl group. Form a triphosphate bond.
4. Oxidation: Oxidation of triphosphate esters to more stable triphosphate esters.
5. Capping: Capping any unreacted 5'-hydroxyl groups to prevent the formation of truncated sequences in subsequent cycles.
6. Cutting and Deprotection: After adding the last nucleotide, the oligonucleotide is cut from the solid carrier and undergoes overall deprotection, removing all protective groups on the base and phosphate backbone.

Key Advantages

• High stability: 2'-O-methyl modification makes RNA less easily degraded by RNase in vivo, and compared to natural nucleic acid molecules, the half-life in vivo can be extended several times to tens of times.
• Strong specificity: The base pairing of DNA units is rigorous and can accurately identify target sequences (such as single nucleotide polymorphisms (SNPs)), reducing binding to non target nucleic acids.
• Low immunogenicity: Modified RNA fragments are not easily recognized by immune molecules such as Toll like receptors (TLRs) in the body, reducing inflammatory reactions and other immune side effects, making them more suitable for in vivo applications.

Breakthrough in Preparation and Purification Technology

Core Services at Creative Biolabs

Equipped with rich expertise in oligonucleotide production and chemical modifications, Creative Biolabs is confident in offering unbeatable synthesis services of RNA and DNA chimeric oligos with 2'-O-Methyl modification for global clients. Our advanced oligonucleotide synthesis platform is composed of nucleic acid synthesizers, nucleic acid purification systems, and nucleic acid analysis instruments, allowing delivery of micrograms or even hundreds of grams oligos in a short time. Besides, we also provide a variety of strategies to improve properties of chimeric oligos, for example, by adding 2'-O-Methyl RNA and phosphorothioate linkages into oligos to further improve their nuclease resistance, by retaining an RNase H activating domain in 2'-O-Methyl RNA chimeric oligos to enable them possess the activity of RNase H cleavage.

Features of Our Services

• An experienced expert team providing real-time technical guidance
• Advanced technical platform feasible for different production scales and purification strategies
• Highly stable oligonucleotide product amenable to different downstream applications and research purposes
• Best after-sale service

Purification Methods for 2'-O-Methyl RNA Bases

The success of any oligonucleotide therapy depends on the purity of the final product. Pollutants, truncated sequences, and adverse byproducts can all affect the effectiveness and safety of candidate drugs. At Creative Biolabs, we use state-of-the-art purification techniques to ensure the highest quality of our 2'-O-methylRNA chimeras. Our main purification method is high-performance liquid chromatography (HPLC), which is the gold standard for separating complex mixtures. HPLC is applied in two key stages:

Frequently Asked Questions

Customer Review

"Our team has long been dedicated to research on regulating hepatitis B virus (HBV) gene expression. Previously, our experiments targeting viral mRNA interference were repeatedly hindered by the fact that conventional oligonucleotides were easily degraded by nucleases in vitro, yet triggered a significant immune response upon in vivo administration. The overall experience, from initial communication to product delivery, exceeded expectations: Creative Biolabs' technical team not only optimized the base length and modification ratio of the chimera based on the target sequence we provided, but also predicted the Tm value and nuclease resistance through simulation experiments, providing invaluable design advice. The chimera we received was accompanied by a detailed quality control report, demonstrating a purity of 97% as determined by HPLC, and the MALDI-TOF mass spectrometry results fully matched the predicted molecular weight."

— Dr. Elena Marquez, Director of Antiviral Research at a Leading Biopharmaceutical Company

Don't Hesitate, Contact Us!

Creative Biolabs has organized an expert team with extensive experience in the field of custom oligonucleotide synthesis and modification. We are committed to providing end-to-end custom oligo modification services from consultation, oligo design, production, and purification, to oligo quality control and delivery. Please don't hesitate to contact us for more details.

Reference

1. Monaco P L, Marcel V, Diaz J J, et al. 2′-O-methylation of ribosomal RNA: towards an epitranscriptomic control of translation? Biomolecules, 2018, 8(4): 106. https://doi.org/10.3390/biom8040106 (Distributed under Open Access license CC BY 4.0, without modification.)