General structure of RNA (2'-O-TBDMS) phosphoramidites
RNA (2'-O-TBDMS) phosphoramidites are the key monomeric building blocks used in solid-phase phosphoramidite chemistry for the synthesis of synthetic RNA oligonucleotides. Unlike DNA synthesis, RNA synthesis presents a fundamental challenge: the highly reactive 2'-hydroxyl group on the ribose ring is prone to side reactions such as branching and degradation during chain assembly. Consequently, RNA monomers must incorporate an orthogonal protecting group at this critical site – protecting the 2'-hydroxyl throughout synthesis and selectively removing it only after the full RNA chain has been assembled, without interfering with the removal of other protecting groups.
TBDMS stands for tert‑butyldimethylsilyl, the most established and widely adopted 2'-O-protecting group in RNA synthesis. A standard RNA (2'-O-TBDMS) phosphoramidite monomer incorporates the following structural features:
RNA-Specific Sugar Backbone – ribose (featuring a 2′-hydroxyl group) replaces the deoxyribose of DNA, with Uridine (U) taking the place of Thymidine (T) alongside the other natural RNA bases (A, G, C).
2'-O-TBDMS Protecting Group – the tert‑butyldimethylsilyl group is attached to the 2'-oxygen, shielding the 2'-hydroxyl throughout the synthesis to prevent side reactions and chain migration/degradation. The TBDMS group is stable to acid and most oxidizing conditions but can be efficiently removed under mild conditions using fluoride ions (e.g., triethylamine trihydrofluoride, TEA·3HF) after full RNA assembly.
Nucleobase Protecting Groups – exocyclic amines on A, G and C are protected with acyl groups (e.g., A: Bz or Ac; G: iBu or Ac; C: Ac or Bz) to prevent side reactions. Fast‑deprotection variants are available for sequences containing sensitive bases.
5'-O-DMT Protecting Group and 3'-Phosphoramidite Group – as with DNA monomers, the 5'-DMT reversibly blocks the 5'-hydroxyl, ensuring that exactly one monomer is added per cycle. The 3'-phosphoramidite group is activated by an activator (e.g., tetrazole) and couples to the free 5'-OH of the growing RNA chain.
On an automated RNA synthesizer, the synthesis proceeds through a four‑step cycle:
Deblocking – dichloroacetic or trichloroacetic acid removes the DMT group from the 5'-terminus, exposing the 5'-OH.
Coupling – the next RNA (2'-O-TBDMS) phosphoramidite and an activator are introduced; the activated amidite couples with the 5'-OH to form a phosphite triester bond.
Capping – any unreacted 5'-OH groups are acetylated (e.g., with acetic anhydride) to prevent deletion (n‑1) impurities.
Oxidation – the unstable phosphite triester is oxidized (typically with iodine) to a stable phosphate diester, forming the natural RNA backbone.
After the cycle is repeated to assemble the full‑length sequence, fluoride ions (e.g., TEA·3HF) are used to remove the 2'-O-TBDMS groups, followed by ammonia to remove the base protecting groups, yielding the final full‑length RNA product.
Hongene's Core Advantages for RNA (2'-O-TBDMS) Phosphoramidites
Complete Product Portfolio – Standard to Specialty Monomers
Hongene offers a comprehensive range of RNA (2'-O-TBDMS) phosphoramidite monomers. Beyond the four standard ribonucleosides (A, G, C, U) with various base protecting groups, the portfolio includes numerous specialty monomers:
Non‑natural base monomers (e.g., Inosine, Pseudouridine, N6‑isopentenyladenosine i6A, and other modified RNA monomers)
Rare / epitranscriptomic-modified monomers.
Representative products include: 5'-O-DMTr-2'-O-TBDMS-L-C(Ac)-3'-CE-Phosphoramidite (Cat. No. PR3-083), 5'-O-DMTr-2'-O-TBDMS-L-U-3'-CE-Phosphoramidite (Cat. No. PR5-064), 5'-O-DMTr-2'-O-TBDMS-L-A(Bz)-3'-CE-Phosphoramidite, and 5'-O-DMTr-2'-O-TBDMS-L-G(iBu)-3--CE-Phosphoramidite, among others.
Ultra‑High Purity & Rigorous Quality Control
Each batch is tested by HPLC/UPLC with HPLC purity ≥98.0% as a standard. Every monomer is accompanied by a full Certificate of Analysis (CoA) covering HPLC purity, moisture content, retention time, mass spectrometry, and impurity profiles. An in‑house impurity reference library enables precise tracking of degradation and process‑related impurities.
GMP Compliance & Large‑Scale Commercial Supply Capability
Hongene‘s manufacturing follows strict GMP standards and has been audited by multiple global pharmaceutical companies. The company operates 58 tons/year phosphoramidite capacity and 54,000 L/year NTP capacity, with a 1.67 million sq. ft oligonucleotide manufacturing facility equipped with multiple OligoPilot™ and OligoProcess™ synthesizers (10–1,800 mmol). 48 flexible production lines enable seamless scale‑up of API production from gram to ton scale, meeting NMPA, FDA, and EMA standards. Hongene was among the world‘s largest suppliers of mRNA vaccine raw materials during the COVID‑19 pandemic, validating its large‑scale reliable delivery capability.
Continuous Innovation & Deep Technical Expertise
With nearly 30 years of industry experience, Hongene has established a secure, reliable supply chain for RNA raw materials. The company‘s proprietary Chemoenzymatic Ligation Platform and other core technology patents, combined with licensing agreements for exNA™ and other novel backbone modification technologies through collaborations with top research institutes such as UMass Medical School, position Hongene to drive next‑generation oligonucleotide therapeutics.
Main Applications of 2'-MOE Phosphoramidites
Life Science Research
Synthesis of siRNA, miRNA, antisense oligonucleotides (ASOs), CRISPR gRNA, and RNA aptamers for gene functional analysis, RNA interference studies, characterization of gene regulatory networks, and structural biology (e.g., X‑ray crystallography and NMR).
RNA Therapeutics & Gene Therapy
Key raw materials for the synthesis of small interfering RNA (siRNA), antisense oligonucleotides (ASOs), aptamers, and CRISPR guide RNA (gRNA). Hongene's GMP‑grade RNA phosphoramidites have supported the raw material supply for multiple clinical‑stage and approved nucleic acid drugs.
mRNA Vaccines & Therapies
Synthesis of DNA templates for in vitro transcription of mRNA and saRNA, as well as direct incorporation of modified nucleotides into mRNA sequences. Hongene‘s products are widely deployed in both R&D and commercial manufacturing of mRNA vaccines and therapeutic mRNAs, including their pivotal role as a core COVID‑19 vaccine raw material supplier.
Synthetic Biology & Functional RNA
Construction of genetic circuits, development of ribozymes and riboswitches, design of artificial RNA nanostructures, and engineering of RNA‑based biosensors.
Epitranscriptomics & RNA Modification Research
Synthesis of RNA sequences containing specific rare/modified bases (e.g., i6A, pseudouridine, 5‑methylcytidine) for precise studies of the roles of RNA modifications in gene expression regulation, translation efficiency, and RNA stability.
