Oligonucleotide synthesis capping. Due to the chemical nature of N7 -methylguanosine, the synthesis of RNAs possessing a cap structure at the 5′ end is still a significant challenge. (top) Sep Pak purified products. When synthetically produced, these strands are an essential part of molecular biology, enabling applications in gene editing, diagnostics, therapeutics, and research. One capping-related impurity has an additional mass of 98 amu from the parent oligonucleotide. With a renewed and growing interest in therapeutic oligonucleotides across the pharmaceutical industry, pressure is increasing on drug developers to take more seriously the sustainability ramifications of this modality. Acetylation of any unreacted 5′-OH groups (“capping”) with acetic anhydride and dimethylaminopyridine as a catalyst. With 12 oligonucleotide drugs reaching the market to date and hundreds more in clinical trials and preclinical development, the current state of the art in oligonucleotide Fig. 4. Oligonucleotide Synthesis Phosphoramidite Method This section explores the detailed chemistry of each of the four steps [step 1 (detritylation), step 2 (coupling), step 3 (capping), and step 4 (oxidation)] in the phosphoramidite method. The ASO field is an emerging area of drug development that targets the disease source at the RNA Oligonucleotide Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, research, and forensics. 3). Such approvals range from antisense oligos (ASOs) to small interfering RNAs (siRNAs) to aptamers, reinforcing the multidisciplinary nature of this class of drugs. It begins with a discussion of an early synthesis method involving a phosphoryl chloride intermediate, which proved unstable and prone to hydrolysis. However, the reaction is also responsible for the formation of a number of impurities. We quantified these synthetic errors by next-generation sequencing and revealed that the quality of the Introduction Our lineup of nucleic acid synthesis reagents focuses on ancillary reagents and dehydration solvents for use in the synthesis of oligonucleotides using the phosphoramidite method. This transformation has been driven by multiple converging Jul 20, 2021 · Different classes of oligonucleotide therapeutics exist, referring to their mode of interaction with the target molecules. Ultramild capping reagent A is a 85:10:5 mixture of THF, pyridine, and phenoxyacetic anhydride, designed for gentle 5’-OH capping in sensitive oligonucleotide synthesis protocols. Syntheses on the 3'-Uaq Cap CPG (1) produce oligonucleotides with a 3’-cap. The length of the oligonucleotide is usually denoted by the term “mer,”which is Greek for “part. Oct 26, 2025 · The meaning of OLIGONUCLEOTIDE is a short nucleic-acid chain usually consisting of up to approximately 20 nucleotides. Caps are covalently appended residues that stabilize double helices by binding to termini. Substitutions, insertions, and deletions derived from synthetic oligonucleotides are the hurdles for the synthesis of long DNA such as genomes. As a result, TGK accepts many variants at the 5'-end, including oligonucleotide primers containing the desired cap. The elongation cycle is repeated until the oligonucleotide is completed (Fig. Oligonucleotide synthesis in brief locking, activation, capping and oxidation. Oligonucleotide syntheses by the phosphotriester method. Jul 20, 2021 · Different classes of oligonucleotide therapeutics exist, referring to their mode of interaction with the target molecules. Robust and cost-efficient oligonucleotide synthesis Intellectual Property Notice: The Biopharma business of GE Healthcare was acquired by Danaher on 31 March 2020 and now operates under the CytivaTM brand. Whereas enzymes synthesize DNA and RNA only in a 5' to 3' direction, chemical Nov 29, 2022 · A comparative assessment of the principal innovations in the synthesis of these molecules is described herein, with a major focus on solid-phase synthesis (SPS), describing particularly the less-explored field of solid-phase oligonucleotide synthesis (SPOS). Glen Research is pleased to offer a new controlled pore glass support for the synthesis of oligonucleotides. Incomplete deblocking causes sequence deletions that are not terminated in the capping step, the resulting failures of which continue to propagate during subsequent coupling cycles. Jul 20, 2023 · The term oligonucleotide is derived from the Greek “oligo,” which means few or small. Sep 22, 2016 · but employs a distinct synthesis procedure termed SOSICS (stereocon-trolled oligonucleotide synthesis with iterative capping and sulfuriza-tion), with a sequence of capping and sulfurization leading to efficient, spontaneous release of the chiral auxiliary Note, Supplementary Figs. Synthesis of Oligonucleotides Automated synthesis of oligonucleotides Advanced synthesis chemistry and state-of-the-art parallel DNA synthesisers allow today for a fast and efficient production of oligonucleotides of almost all sequences. This transformation has been driven by multiple converging Apr 14, 2023 · Oligos, short for oligonucleotide, are short synthetic strands of DNA or RNA. Apr 14, 2023 · Oligos, short for oligonucleotide, are short synthetic strands of DNA or RNA. The technique is extremely useful in current laboratory practice because it provides a rapid and inexpensive access to custom-made oligonucleotides of the desired sequence. Feb 21, 2010 · Traditional methods of solid-phase oligonucleotide synthesis (3, 4), where each oligonucleotide is synthesized individually on a separate column or in a multi-well plate, produce high yields but are costly. Solid phase synthesis allows for automation and purification after each step. For the assembly of long genes or even whole genomic sequences, the cost of the starting oligonucleotides alone can be prohibitive. Oligonucleotide synthesis outside of biological systems is the chemical synthesis of nucleic acids of a predetermined sequence that may contain nucleosides beyond those that are naturally occurring. In the present work, we combined a chemical synthesis method and an enzymatic methylation assay in order to produce large amounts of RNA oligonucleotides carrying a cap-0 or cap-1. Let’s look in greater detail at what they are, how they are used, and why they are important for genetic research. Oligo synthesis is made following a synthesis cycle of 4 reactions : Deblocking of the nucleotide, coupling of nucleotide, capping and oxidation. Synthetic DNA of defined sequences are commonly termed “oligonucleotides,” which are primarily composed of four different types of nucleosides linked through well-defined deoxyribose phosphate. Figure 2 displays the monomeric units that are necessary in this procedure. Oligonucleotides binding to mRNA rely on a Watson-Crick base-paring mechanism. The impurity has been found to arise by conversion of a protected guanine nucleobase to N (2)-acetyl-2,6-diaminopurine. After the entire oligonucleotide is built, it is cleaved from its solid support, deprotected, and furt Cap A - 20% NMI in Acetonitrile Used to 'cap' unreacted OH groups during oligo synthesis cycle, to prevent unwanted side reactions. The efficiency of the coupling cycle is critical to the success of a synthesis. Oxidation of the phosphite to phosphate with iodine/water. There have been 20 oligonucleotide drugs approved by the FDA since 1998, and it's climbed to roughly two new drugs every year since 2016. The n+98 amu 3. Although powerful, this approach suffers from fundamental limitations that make it unsuitable for large-scale manufacturing. ” There have been 20 oligonucleotide drugs approved by the FDA since 1998, and it's climbed to roughly two new drugs every year since 2016. Over the past ten years, because of refinement in synthesis In contrast to most other RNA polymerases used for conventional in vitro transcription, primer-dependent RNA synthesis does not require the use of pppG to initiate RNA synthesis. This article deeply explores the core role of phosphoramidite chemistry in oligonucleotide synthesis, and systematically analyzes the action mechanism of key raw materials such as base-protected phosphoramidites and dimer phosphoramidites and their advantages in improving synthesis efficiency, purity and sequence fidelity. Antisense oligonucleotides (ASOs) bind sequence specifically to the target RNA and modulate protein expression through several different mechanisms. An oligonucleotide is a short chain of nucleotides (nitrogen-containing units linked to a sugar and a phosphate group) that consist of either single- or double-stranded DNA or RNA. 1,19 mer showing a promi- nent n-l component; 2, failed synthesis of 19 mer with a 3’-purine Jun 15, 2023 · Traditional approaches to oligonucleotide synthesis rely on solid-phase phosphoramidite chemistry, which involves iterative coupling, capping, oxidation, and deprotection steps for each nucleotide addition (6). Unless otherwise stated, syntheses did not include a capping step. 1 and (Fig. Biosearch Technologies ofers oligonucleotide synthesis instruments that support production at a range of scales, flexibility, and throughputs. Aug 27, 2024 · Abstract Chemical synthesis of phosphoromonothioate oligonucleotides (PS-ONs) is not stereo-specific and produces a mixture of Rp and Sp diastereomers, whose disparate reactivity can complicate applications. Once confined to PCR primers and hybridization probes, oligos now play central roles in developing novel therapeutics, powering genome-editing technologies and enabling rapid diagnostics. 1-3 Caps increase duplex melting points and improve base pairing fidelity by selectively binding to Watson-Crick base Aug 1, 1996 · Introduction High product yields from oligonucleotide synthesis require quantitative removal of the dimethoxytrityl (DMT) group during the deblocking step. We provide reagents of suitable quality for nucleic acid synthesis by leveraging our unique liquid preparation, synthesis, dehydration, and analysis technologies developed over years of reagent The invention includes a method for capping failure sequences in oligonucleotide synthesis, and methods of synthesizing oligonucleotides which include the capping method of the invention as a step. There is increasing evidence (1, 2) that the deblocking Oligonucleotide synthesis is the chemical synthesis of relatively short fragments of nucleic acids with defined chemical structure (sequence). Although the current methods to separate these diastereomers which rely on chromatography are constantly improving, many Rp and Sp diastereomers are still co-eluted. You are here: Oligonucleotide Synthesis Reagents Solvents and Reagents Capping Solutions Capping A Aug 1, 2014 · The acetyl 'capping' reaction routinely employed during phosphorothioate oligonucleotide synthesis has been implicated in the formation of an impurity species with a mass 41 amu greater than the expected oligonucleotide molecule. Synthesis The overall procedure for obtaining these oligonucleotides is illustrated in Figure 1 and follows the stepwise process: automated Solid-phase Synthesis → Deprotection → Purification → Quantification → Characterization. Nov 21, 2024 · This review examines several oligonucleotide synthesis techniques, including phosphodiester, phosphotriester, and phosphoramidite approaches. Oct 29, 2025 · Oligonucleotides – short, synthetic sequences of DNA or RNA – have evolved from basic lab reagents to powerful enablers of precision medicine. PAGE-autoradiography of various reaction products. Certain collateral materials (such as application notes, scientific posters, and white papers) were created prior to the Danaher acquisition and contain various GE owned trademarks and font Aug 26, 2024 · What is oligonucleotide synthesis? Oligonucleotide synthesis is the chemical process used to create short strands of nucleic acids, such as DNA or RNA, with a specific sequence. Samples were chosen to show the various types of reaction mixtures encoun- tered. Capping reagent B1 (40% acetic anhydride in acetonitrile) for ÄKTA OligoPilot. In a so-called solid-phase synthesis the desired oligonucleotide sequence is synthesised onto an inert solid support, from which it is cleaved after the Jun 18, 2024 · Controlled enzymatic synthesis Controlled enzymatic synthesis is a method for the de novo production of oligonucleotides that combines elements of solid-phase synthesis and of enzymatic Aug 1, 2016 · The acetyl capping reaction used throughout solid phase oligonucleotide synthesis is meant to minimize n−1 deletionmer impurities by terminating sequences that fail to couple to a phosphoramidite. Jul 1, 2025 · The structure of an oligonucleotide is similar to that of natural nucleic acids: it consists of a sugar-phosphate backbone with attached nitrogenous bases (adenine, cytosine, guanine, thymine in DNA, and uracil instead of thymine in RNA). Reliable capping solution tailored for phosphoramidite oligo synthesis systems. 1b; Supplementary. Oligonucleotide Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, research, and forensics. Typical oligonucleotide synthesis using nucleoside phosphoramidites occurs in a four-step process: deblocking, coupling, capping, and oxidation. Oligonucleotide Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications in genetic testing, research, and forensics. An oligonucleotide is defined as a short DNA or RNA molecule, either single- or double-stranded, which includes antisense oligonucleotides, RNA interference, and aptamer RNAs. Here, based on sulfur The phosphoramidite method is commonly used, involving cycles of deprotection, coupling of the next nucleotide, oxidation to stabilize the linkage, and capping. hsd mqqf drb dfe lz33 5mo rb4b ihlwm 4l mn6ob