Long oligodeoxynucleotides: chemical synthesis, isolation via catching-by-polymerization, verification via sequencing, and gene expression demonstration

• Yipeng Yin,
• Reed Arneson,
• Alexander Apostle,
• Adikari M. D. N. Eriyagama,
• Komal Chillar,
• Emma Burke,
• Martina Jahfetson,
• Yinan Yuan and
• Shiyue Fang

Beilstein J. Org. Chem. 2023, 19, 1957–1965, doi:10.3762/bjoc.19.146

• ODN sequences are provided in Supporting Information File 1) GFP gene construct was divided into a 399 and 401 nt ODNs for automated synthesis (step 1, Figure 1). The syntheses were carried out in commercial 0.2 µmol 2000 Å CPG columns on an ABI 394 DNA/RNA synthesizer using phosphoramidite chemistry

• caused by long ODNs, the loading of the synthesis column was reduced from 0.2 µmol to 0.1 µmol. This was achieved by applying a solution containing 0.05 M 5'-DMTr-dT-phosphoramidite (1a, Scheme 1) and 0.05 M 5'-Bz-dT-phosphoramidite (1b) for the coupling step in the first synthetic cycle. Because the Bz

• cycle, the polymerizable tagging phosphoramidite 2, of which the synthesis was described earlier [20] (Scheme 1), was used in the coupling step, and the reaction was repeated three times for 5 minutes each time to maximize the yield of tagging the full-length sequences with the polymerizable

Synthesis of ether lipids: natural compounds and analogues

• Marco Antônio G. B. Gomes,
• Alicia Bauduin,
• Chloé Le Roux,
• Romain Fouinneteau,
• Wilfried Berthe,
• Mathieu Berchel,
• Hélène Couthon and
• Paul-Alain Jaffrès

Beilstein J. Org. Chem. 2023, 19, 1299–1369, doi:10.3762/bjoc.19.96

• method was developed for the incorporation of the phosphocholine polar head group that makes use of the phosphoramidite 8.2 which is a weakly air sensitive reagent [85]. This method was applied for the synthesis of PAF as illustrated in Figure 8 [86]. The alcohol 8.1 reacted with 8.2 in the presence of

• ) produced 20.4. Then, the incorporation of the phosphoinositol moiety was achieved by using phosphoramidite chemistry. First, the alcohol 20.4 reacted with O-benzyl-N,N,N’,N’-tetraisopropylphosphorodiamidite to produce the phosphoramidite 20.5 that subsequently reacted with dibenzyl-tris(dibenzylphosphate

Enolates ambushed – asymmetric tandem conjugate addition and subsequent enolate trapping with conventional and less traditional electrophiles

• Péter Kisszékelyi and
• Radovan Šebesta

Beilstein J. Org. Chem. 2023, 19, 593–634, doi:10.3762/bjoc.19.44

• (phosphite/phosphine-pyridine amide, phosphine-sulfoxide, phosphoramidite, MINBOL, see Figure 1) and they usually showed excellent diastereoselectivity (dr >20:1). The catalytic systems even with low catalyst loadings tolerated both electron-donating and withdrawing groups on the aromatic substituents

• the trapping of metal enolates by Michael reactions with nitroalkenes 122 and disulfonyl ethylenes 124 (Scheme 31) [67]. The conjugate additions of dialkylzinc, Grignard, and trialkylaluminum reagents to cyclic enones 121 were realized using previously established chiral phosphoramidite, carbene or

• in the first step by the phosphoramidite ligand L39 (92% ee, dr 1:1) (Scheme 60). Recently, Liu and co-workers reported the stereoselective synthesis of the tricyclic core of dodecahydrodibenzo[b,d]furan skeleton containing 12-epi-JBIR-23 and -24 [111]. Besides their intriguing complex structures

Heteroleptic metallosupramolecular aggregates/complexation for supramolecular catalysis

• Prodip Howlader and
• Michael Schmittel

Beilstein J. Org. Chem. 2022, 18, 597–630, doi:10.3762/bjoc.18.62

• demonstrated by Reek and co-workers by using a previously reported heteroleptic bisporphyrin cage [76]. The tetragonal prismatic nanocage 47 consisted of two zinc-porphyrin units along the two tetragonal faces (Figure 11), which allowed encapsulation of the chiral phosphoramidite 48 as a precursor for the

The role of chemistry in the success of oligonucleotides as therapeutics

• Pawan Kumar and
• Tom Brown

Beilstein J. Org. Chem. 2022, 18, 197–199, doi:10.3762/bjoc.18.22

• quantities of oligonucleotides it would not have been possible to develop them as therapeutic agents. Forty years ago, the synthesis of an oligonucleotide in the lab was a huge task. Since then, seminal work from the Caruthers lab has solved this problem with the introduction of phosphoramidite chemistry [4

• ]. This proved an outstanding breakthrough and is currently catalyzing the development of many new technologies including next generation siRNAs, antisense oligonucleotides, and CRISPR-based gene editing systems. Thanks to this phosphoramidite approach, it has also been possible to mass-produce

Cationic oligonucleotide derivatives and conjugates: A favorable approach for enhanced DNA and RNA targeting oligonucleotides

• Mathias B. Danielsen and
• Jesper Wengel

Beilstein J. Org. Chem. 2021, 17, 1828–1848, doi:10.3762/bjoc.17.125

• between the modified nucleobase and the corresponding guanidine, which resulted in an increase in Tm of 16 °C, i.e., in the same range as obtained with the original G-clamp (Table 3A) [59]. Generally, conversions of nucleoside phosphoramidite synthons have been explored only rarely. However, the

• commercially available 3’-phosphoramidite derivative of 5’-O-dimethoxytrityl-2’-O-methyluridine could be converted into an N4-triazole-modified 2’-OMe-cytidine phosphoramidite [60]. This concept was later used to prepare spermine-functionalized 2’-OMe and 2’-O-((2-methoxy)ethyl) (MOE)cytidine phosphoramidites

• be taken when conjugating amine moieties onto guanine. Interestingly, in a post-oligo synthetic modification approach, a 2-fluoro-6-p-nitrophenylethyl-2’-deoxyinosine-3'-phosphoramidite monomer was incorporated twice into an 11-mer ON, whereupon spermine was attached to the 2-position of the purine

Double-headed nucleosides: Synthesis and applications

• Vineet Verma,
• Jyotirmoy Maity,
• Vipin K. Maikhuri,
• Ritika Sharma,
• Himal K. Ganguly and
• Ashok K. Prasad

Beilstein J. Org. Chem. 2021, 17, 1392–1439, doi:10.3762/bjoc.17.98

• double-headed nucleosides 4a,b were dimethoxytritylated (DMTr), phosphitylated, and incorporated into DNA oligonucleotides using the standard automated phosphoramidite method. The UV-based melting temperature (Tm) of hybrids of the modified oligonucleotides with complementary DNA strands were studied

• nucleoside 37 (Scheme 9) [34]. The double-headed nucleoside 37 was dimethoxytritylated, phosphitylated, and incorporated into 11- to 13-mer oligonucleotides using the standard automated phosphoramidite method. The UV-based melting temperature (Tm) of hybrids of the modified oligonucleotides with

• nucleoside monomers 125a–c were incorporated into oligodeoxyribonucleotides via phosphoramidite derivatization of the C-3′ hydroxy group present in the moiety. The oligonucleotides thus synthesized were found to stabilize the duplex formed with complementary DNA [69]. Nielsen and co-workers [43][70

Recent advances in palladium-catalysed asymmetric 1,4–additions of arylboronic acids to conjugated enones and chromones

• Jan Bartáček,
• Jan Svoboda,
• Martin Kocúrik,
• Jaroslav Pochobradský,
• Alexander Čegan,
• Miloš Sedlák and
• Jiří Váňa

Beilstein J. Org. Chem. 2021, 17, 1048–1085, doi:10.3762/bjoc.17.84

• the reaction (Scheme 6) [40]. A different approach using microwave irradiation was explored by the group of Toma et al. [41]. After an initial tuning of the reaction conditions of a catalytic system based on Pd(OAc)2/2,2’-bipy several optically pure phosphoramidite and diphosphine ligands in

Beyond ribose and phosphate: Selected nucleic acid modifications for structure–function investigations and therapeutic applications

• Christopher Liczner,
• Kieran Duke,
• Gabrielle Juneau,
• Martin Egli and
• Christopher J. Wilds

Beilstein J. Org. Chem. 2021, 17, 908–931, doi:10.3762/bjoc.17.76

• synthesize the nucleoside dimer phosphoramidite with the appropriate amide linkage, which can then be introduced into the strand by solid-phase synthesis. These dimers are synthesized by using an amide coupling reagent to condense a 3'-carboxylic acid nucleoside with a 5'-amine nucleoside, where the

• obtained through condensation with N,N-dicyclohexylcarbodiimide (DCC) giving rise to homopolymeric tetramers of either G-GNA or T-GNA [97]. In 1996, Acevedo and Andrews were the first to demonstrate the synthesis of GNA nucleoside phosphoramidite derivatives as well as the ability of the phosphoramidite

• -di-O-benzoyl-2-deoxy-2-fluoro-α-ᴅ-arabinofuranosyl bromide (Scheme 7) [176][177][178][179][180]. Both β-ᴅ-arabino and 2'-fluoro-β-ᴅ-arabinofuranose nucleosides can be converted to phosphoramidite derivatives for incorporation into oligonucleotides for solid-phase synthesis [178][181][182][183][184

DNA with zwitterionic and negatively charged phosphate modifications: Formation of DNA triplexes, duplexes and cell uptake studies

• Yongdong Su,
• Maitsetseg Bayarjargal,
• Tracy K. Hale and
• Vyacheslav V. Filichev

Beilstein J. Org. Chem. 2021, 17, 749–761, doi:10.3762/bjoc.17.65

• -phase DNA or RNA synthesis, the introduction of SaRNA monomers into an RNA backbone involved a 14-step preparation of the phosphoramidite for the SaRNA-TT dinucleotide. Similarly, the incorporation of BCNS groups on a DNA backbone requires the synthesis of thymidine and 2’-OMe-uridine phosphoramidites

Synthesis and properties of oligonucleotides modified with an N-methylguanidine-bridged nucleic acid (GuNA[Me]) bearing adenine, guanine, or 5-methylcytosine nucleobases

• Naohiro Horie,
• Takao Yamaguchi,
• Shinji Kumagai and
• Satoshi Obika

Beilstein J. Org. Chem. 2021, 17, 622–629, doi:10.3762/bjoc.17.54

• for the guanidine moiety because it can be easily removed under the basic conditions (ammonia/methylamine solution) used for the DNA synthesis [20]. The phosphoramidite synthesis was started from 2'-amino-LNAs 1a–c, which were rapidly prepared via the transglycosylations of 2'-amino-LNA-T [25]. First

• modified with GuNA[Me]-A, -G, or -mC (0.2 µmol scale) was performed using the nS-8 oligonucleotide synthesizer (GeneDesign, Inc.) according to the standard phosphoramidite protocol with 0.5 M 5-ethylthiotetrazole as an activator. The protocol is similar to that described in reference [20]. A Custom Primer

Incorporation of a metal-mediated base pair into an ATP aptamer – using silver(I) ions to modulate aptamer function

• Marius H. Heddinga and
• Jens Müller

Beilstein J. Org. Chem. 2020, 16, 2870–2879, doi:10.3762/bjoc.16.236

• phosphoramidite-protected imidazole nucleoside as required for automated DNA solid-phase synthesis was synthesized as previously reported [30]. The oligonucleotides were synthesized on a DNA/RNA synthesizer H-8 (K&A Laborgeräte) using standard protocols for automated solid-phase synthesis (coupling time: 1000 s

• for unnatural nucleosides). All natural nucleosides (A, T, G, and C), the fluorescein phosphoramidite as well as the DABCYL-CPGs were obtained from Glen Research. The oligonucleotides were deprotected and cleaved from the support by incubation in AMA solution (1:1, 40% methylamine and aqueous ammonia

Changed reactivity of secondary hydroxy groups in C8-modified adenosine – lessons learned from silylation

• Jennifer Frommer and
• Sabine Müller

Beilstein J. Org. Chem. 2020, 16, 2854–2861, doi:10.3762/bjoc.16.234

• , functionalized phosphoramidite building blocks of all four nucleosides are highly desired. The number of commercially available RNA phosphoramidites that carry a suitable functionality for post-synthetic attachment of dyes, reporter groups or other conjugates is still rather limited. In particular, monomer

• synthesized a number of pyrimidine and purine derivatives carrying amino linkers of different length and flexibility [13][20]. Linker-modified uridine derivatives, upon conversion into phosphoramidite building blocks, were incorporated in RNA and used for a systematic study of distance determination of

• phosphoramidite building block 9 (Scheme 2). Results and Discussion Typically, the synthesis of C8-alkynyl derivatives relies on C8-bromoadenosine as reactant for the Sonogashira cross-coupling reaction to introduce the amino linker N-(propyn-2-yl)-6-(trifluoroacetamido)hexanamide (L) bearing an alkynyl moiety

Nonenzymatic synthesis of anomerically pure, mannosyl-based molecular probes for scramblase identification studies

• Giovanni Picca,
• Markus Probst,
• Simon M. Langenegger,
• Oleg Khorev,
• Peter Bütikofer,
• Anant K. Menon and
• Robert Häner

Beilstein J. Org. Chem. 2020, 16, 1732–1739, doi:10.3762/bjoc.16.145

• citronellol derivative via a phosphodiester bridge. Moreover, a novel phosphoramidite chemistry-based method offers a straightforward approach for the non-enzymatic incorporation of the saccharide moiety in an anomerically pure form. Keywords: carbohydrates; citronellol; phosphoramidite; photoclickable

• into stereodefined, anomeric phosphoramidite derivatives. The method is based on adapted procedures developed for DNA solid-phase synthesis [17]. Phosphoramidite chemistry allows the connection of two molecular entities via a phosphodiester linkage and was found to be perfectly suited for this purpose

• . Reports of using phosphoramidite chemistry for the preparation of carbohydrates via the anomeric position are relatively rare [18][19][20][21][22][23]. Alternatively, the H-phosphonate approach has been used to convert carbohydrates into phosphate-linked derivatives at the anomeric center [24][25][26][27

Copper-catalysed alkylation of heterocyclic acceptors with organometallic reagents

• Yafei Guo and
• Syuzanna R. Harutyunyan

Beilstein J. Org. Chem. 2020, 16, 1006–1021, doi:10.3762/bjoc.16.90

• copper-catalysed asymmetric conjugate addition (ACA) of dialkylzinc reagents to N-substituted 2,3-dehydro-4-piperidones 1 in order to access useful chiral piperidine derivatives (Scheme 1A) [15]. They found the catalytic system based on the chiral phosphoramidite L1 and a copper salt to be the most

• an N-acyl-4-methoxypyridinium salt using a copper/phosphoramidite catalytic system (Scheme 3) [20][21]. The authors highlighted that the N-acylpyridinium salts were unstable species and that their instability affected the regioselectivity of the dearomatisation process upon the nucleophilic addition

• acceptors with trialkylaluminium reagents was explored by the group of Alexakis in 2009 (Scheme 11) [41]. Various chiral phosphoramidite ligands, in combination with a copper salt, were found to be efficient catalysts for this transformation, with the best results obtained with the ligand L16. Feringa and

Recent advances in Cu-catalyzed C(sp³)–Si and C(sp³)–B bond formation

• Balaram S. Takale,
• Ruchita R. Thakore,
• Elham Etemadi-Davan and
• Bruce H. Lipshutz

Beilstein J. Org. Chem. 2020, 16, 691–737, doi:10.3762/bjoc.16.67

• alcohols (Scheme 44) [82]. In most cases, high chemical yields along with high ees were obtained when phosphoramidite ligand L28 was used. A wide variety of compounds was prepared, including one with a bisquarternary center (267), although the Bpin residue within it could not be oxidized to the desired

Copper-catalyzed enantioselective conjugate addition of organometallic reagents to challenging Michael acceptors

• Delphine Pichon,
• Jennifer Morvan,
• Christophe Crévisy and
• Marc Mauduit

Beilstein J. Org. Chem. 2020, 16, 212–232, doi:10.3762/bjoc.16.24

• reagents was reported by Alexakis and co-workers in 2010 [14]. After screening various chiral phosphine-based ligands, the combinations of either phosphoramidite L1 with Cu(OTf)2, or (R)-BINAP (L2) with copper thiophenecarboxylate (CuTC) appeared to be the most efficient for the addition of Et2Zn to a

• copper-catalyzed ECA of alkylzirconium reagents to α,β-unsaturated thioesters [33]. Starting from diversely functionalized alkenes, the resulting hydrozirconated adducts were reacted with various β-substituted Michael acceptors in the presence of CuCl and the chiral phosphoramidite L11 (Scheme 12

• addition of diethylzinc to crotonyl N-acyloxazolidinone [48]. The 1,4-product was also formed in high enantioselectivity (>98% ee) and high yield (91%). The same year, Pineschi et al. evaluated various α,β-unsaturated acyl derivatives for the copper/(R,S,S)-phosphoramidite L16-catalyzed addition of

Electrophilic oligodeoxynucleotide synthesis using dM-Dmoc for amino protection

• Shahien Shahsavari,
• Dhananjani N. A. M. Eriyagama,
• Bhaskar Halami,
• Vagarshak Begoyan,
• Marina Tanasova,
• Jinsen Chen and
• Shiyue Fang

Beilstein J. Org. Chem. 2019, 15, 1116–1128, doi:10.3762/bjoc.15.108

• removed with an acid in each synthetic cycle. The exo-amino groups of nucleosides dA, dC and dG are protected with acyl groups, the nascent ODN is anchored to a solid support via a base- or nucleophile-cleavable linker, and in the most widely used phosphoramidite technology the phosphate groups are

• electrophilic ODN synthesis technology, the phosphoramidite monomers 3a–c and the linker 4 (Figure 1) were needed. The construction of linker 4 was reported previously [40]. The synthesis of 3a–c is shown in Scheme 2. The reagent 5 needed for protecting the exo-amino groups of nucleobases was prepared in two

• steps from 1,3-dithiane (6) according to a procedure we reported previously [44]. The dC phosphoramidite monomer 3a was synthesized from compound 9 [45]. The formation of the hindered O-tert-alkyl N-arylcarbamate 10 was found highly challenging [44][46][47]. We tried many conditions and finally found

Syntheses and chemical properties of β-nicotinamide riboside and its analogues and derivatives

• Mikhail V. Makarov and
• Marie E. Migaud

Beilstein J. Org. Chem. 2019, 15, 401–430, doi:10.3762/bjoc.15.36

Synthesis, biophysical properties, and RNase H activity of 6’-difluoro[4.3.0]bicyclo-DNA

• Sibylle Frei,
• Adam K. Katolik and
• Christian J. Leumann

Beilstein J. Org. Chem. 2019, 15, 79–88, doi:10.3762/bjoc.15.9

• -DNA (6’-diF-bc4,3-DNA). The difluorinated thymidine phosphoramidite building block was synthesized starting from an already known gem-difluorinated tricyclic glycal. This tricyclic siloxydifluorocyclopropane was converted into the [4.3.0]bicyclic nucleoside via cyclopropane ring-opening through the

• addition of an electrophilic iodine during the nucleosidation step followed by reduction. The gem-difluorinated bicyclic nucleoside was then converted into the corresponding phosphoramidite building block which was incorporated into oligonucleotides. Thermal denaturation experiments of these

• analog by RNase H. In addition, the RNase H experiment was also performed with the previously report 6’F-bc4,3-DNA (Figure 1) [37]. Results and Discussion Synthesis of the phosphoramidite building block In the literature there exist several procedures to construct an α,α-difluoroketone from a

6’-Fluoro[4.3.0]bicyclo nucleic acid: synthesis, biophysical properties and molecular dynamics simulations

• Sibylle Frei,
• Andrei Istrate and
• Christian J. Leumann

Beilstein J. Org. Chem. 2018, 14, 3088–3097, doi:10.3762/bjoc.14.288

• unsaturated 6’-fluoro[4.3.0]bicyclo nucleotides (6’F-bc4,3-DNA). Two 6’F-bc4,3 phosphoramidite building blocks (T and C) were synthesized starting from a previously described [3.3.0]bicyclic sugar. The conversion of this sugar to a gem-difluorinated tricyclic intermediate via difluorocarbene addition followed

• phosphoramidite building blocks into oligonucleotides was achieved with tert-butyl hydroperoxide as oxidation agent. Thermal melting experiments of the modified duplexes disclosed a destabilizing effect versus DNA and RNA complements, but with a lesser degree of destabilization versus complementary DNA (ΔTm/mod

• duplexes. Results and Discussion Synthesis of the phosphoramidite building blocks Our strategy for the construction of the two phosphoramidite building blocks 10 and 16 envisaged as a key step the formation of a [4.3.0]bicyclic fluoroenone from a tricyclic siloxydifluorocyclopropane through a ring

Hydroarylations by cobalt-catalyzed C–H activation

• Rajagopal Santhoshkumar and
• Chien-Hong Cheng

Beilstein J. Org. Chem. 2018, 14, 2266–2288, doi:10.3762/bjoc.14.202

• the presence of Co(acac)3 and phosphoramidite ligand L afforded alkylated products 33 in high yields and enantioselectivity. A wide range of indoles and styrenes were well tolerated to form the corresponding alkyl products 33 in a branched selective manner. In addition to the directing group

Artificial bioconjugates with naturally occurring linkages: the use of phosphodiester

• Takao Shoji,
• Hiroki Fukutomi,
• Yohei Okada and
• Kazuhiro Chiba

Beilstein J. Org. Chem. 2018, 14, 1946–1955, doi:10.3762/bjoc.14.169

• activated 5’-terminus without difficulty (Scheme S2, Supporting Information File 1). Furthermore, to our delight, the conjugation was compatible with carboxylic acids that are potential nucleophiles for the activated 5’-terminus and can also induce side reactions of the phosphoramidite (Scheme 1). This

Phosphoramidite building blocks with protected nitroxides for the synthesis of spin-labeled DNA and RNA

• Timo Weinrich,
• Eva A. Jaumann,
• Ute M. Scheffer,
• Thomas F. Prisner and
• Michael W. Göbel

Beilstein J. Org. Chem. 2018, 14, 1563–1569, doi:10.3762/bjoc.14.133

• required to assemble oligonucleotides by phosphoramidite chemistry and to ligate them enzymatically, are known to partially degrade nitroxides. This problem can be reduced by postsynthetic introduction of the spin label [13][14][15][16][17][18][19][20][21][22][23][24]. Starting from convertible nucleotides

• oligonucleotides by phosphoramidite building blocks [27][28][29][30][31][32][33][34][35][36][37][38][39][40]. However, some degradation of the spin labels inevitably will occur. Nitroxide labeled DNA strands of the general structures 2 and 4 have been prepared by this way [30][34][37]. In recent years, we have

• therefore developed a third strategy based on photolabile protective groups [41][42][43] finally leading to phosphoramidite 6. This building block behaves in solid-phase RNA synthesis as any normal TOM protected amidite. It is completely stable against the conditions used for strand assembly, RNA

Oligonucleotide analogues with cationic backbone linkages

• Melissa Meng and
• Christian Ducho

Beilstein J. Org. Chem. 2018, 14, 1293–1308, doi:10.3762/bjoc.14.111

• supra). We have reported that partially zwitterionic NAA-modified DNA oligonucleotides can be obtained by standard solid phase-supported automated DNA synthesis if 'dimeric' phosphoramidite building blocks 49 and 50 are employed (Scheme 5) [72][73]. For the synthesis of 'dimeric' phosphoramidites 49 and

• cationic oligomers, i.e., oligonucleotide analogues with the cationic NAA-modification as their sole internucleotide linkage. The phosphoramidite-based synthetic strategy depicted in Scheme 5 was not suitable to reach this goal as it furnishes phosphate diester linkages at least at every second position

• accessible by means of chemical synthesis, which is either based on the application of H-phosphonate (for i) or phosphoramidite-based (for iv) DNA synthesis, or on a massively modified version of DNA synthesis (for ii and iii), or on solid phase-supported peptide synthesis (for iv). Studies on the properties