Synthesis, binding and cellular uptake properties of oligodeoxynucleotides containing cationic bicyclo-thymidine residues

The synthesis and incorporation into oligodeoxynucleotides of two novel derivatives of bicyclothymidine carrying a cationic diaminopropyl or lysine unit in the C(6′)-β position is described. Compared to unmodified DNA these oligonucleotides show Tm-neutral behavior when paired against complementary DNA and are destabilizing when paired against RNA. Unaided uptake experiments of a decamer containing five lys-bcT units into HeLa and HEK293T cells showed substantial internalization with mostly cytosolic distribution which was not observed in the case of an unmodified control oligonucleotide

Synthesis, Pairing, and Cellular Uptake Properties of C(6′)-Functionalized Tricyclo-DNA

Tricyclo-DNA (tc-DNA) is a promising candidate for oligonucleotide-based therapeutic applications exhibiting increased affinity to RNA and increased resistance to nucleases. However, as many other oligonucleotide analogs, tc-DNA does not readily cross cell membranes. We wished to address this issue by preparing a prodrug of tc-DNA containing a metabolically labile group at C(6′) that promotes cellular uptake. Two monomeric nucleoside building blocks bearing an ester function at C(6′) (tc^ee-T and tc^hd-T) were synthesized starting from a known C(6′) functionalized bicyclic sugar unit to which the cyclopropane ring was introduced via carbene addition. NIS-mediated nucleosidation of the corresponding glycal with in situ persilylated thymine afforded the β-iodonucleoside exclusively that was dehalogenated via radical reduction. Diversity in the ester function was obtained by hydrolysis and reesterification. The two nucleosides were subsequently incorporated into DNA or tc-DNA by standard phosphoramidite chemistry. The reactivity of the ester function during oligonucleotide deprotection was explored and the corresponding C(6′) amide, carboxylic acid, or unchanged ester functions were obtained, depending on the deprotection conditions. Compared to unmodified DNA, these tc-DNA derivatives increased the stability of duplexes investigated with Δ<i>T</i>_m/mod of +0.4 to +2.0 °C. The only destabilizing residue was tc^hd-T, most likely due to self-aggregation of the lipophilic side chains in the single stranded oligonucleotide. A decamer containing five tc^hd-T residues was readily taken up by HeLa and HEK 293T cells without the use of a transfection agent

Synthesis, Pairing, and Cellular Uptake Properties of C(6′)-Functionalized Tricyclo-DNA

Tricyclo-DNA (tc-DNA) is a promising candidate for oligonucleotide-based therapeutic applications exhibiting increased affinity to RNA and increased resistance to nucleases. However, as many other oligonucleotide analogs, tc-DNA does not readily cross cell membranes. We wished to address this issue by preparing a prodrug of tc-DNA containing a metabolically labile group at C(6′) that promotes cellular uptake. Two monomeric nucleoside building blocks bearing an ester function at C(6′) (tc^ee-T and tc^hd-T) were synthesized starting from a known C(6′) functionalized bicyclic sugar unit to which the cyclopropane ring was introduced via carbene addition. NIS-mediated nucleosidation of the corresponding glycal with in situ persilylated thymine afforded the β-iodonucleoside exclusively that was dehalogenated via radical reduction. Diversity in the ester function was obtained by hydrolysis and reesterification. The two nucleosides were subsequently incorporated into DNA or tc-DNA by standard phosphoramidite chemistry. The reactivity of the ester function during oligonucleotide deprotection was explored and the corresponding C(6′) amide, carboxylic acid, or unchanged ester functions were obtained, depending on the deprotection conditions. Compared to unmodified DNA, these tc-DNA derivatives increased the stability of duplexes investigated with Δ<i>T</i>_m/mod of +0.4 to +2.0 °C. The only destabilizing residue was tc^hd-T, most likely due to self-aggregation of the lipophilic side chains in the single stranded oligonucleotide. A decamer containing five tc^hd-T residues was readily taken up by HeLa and HEK 293T cells without the use of a transfection agent

2‑Pyrenyl-DNA: Synthesis, Pairing, and Fluorescence Properties

Multiple 2-pyrenyl-C-nucleosides were incorporated into the center of a DNA duplex resulting in stable pyrene self-recognition and excimer formation. This helical pyrene array may find use in DNA-mediated charge transfer and in the creation of DNA-based sensors

Low cost DNA data storage using photolithographic synthesis and advanced information reconstruction and error correction

The current bottleneck for DNA data storage systems is the cost and speed of synthesis. Here, the authors use inexpensive, massively parallel light-directed synthesis and correct for a high error rate with a pipeline of encoding and reconstruction algorithms

An open-source advanced maskless synthesizer for light-directed chemical synthesis of large nucleic acid libraries and microarrays

Large- to ultra-large-scale synthesis of nucleic acids is becoming an increasingly important tool for understanding and manipulating biological systems, as well as for developing new technologies based on engineered biological materials, including DNA-based nanofabrication, aptamers and writing digital data at the molecular level. Several technologies for large-scale synthesis have been developed over the years, but all of them remain inaccessible to most researchers due to their complexity and/or use of proprietary technologies. Here, we present a fully open source, benchtop device for ultra-large scale nucleic acid synthesis that is also highly flexible and adaptable, able to accommodate a wide range of monomers and chemistries, while providing unrestricted access to synthesis parameter space

Defining the Sphagnum Core Microbiome across the North American Continent Reveals a Central Role for Diazotrophic Methanotrophs in the Nitrogen and Carbon Cycles of Boreal Peatland Ecosystems

Nitrogen availability frequently limits photosynthetic production in Sphagnum moss-dominated high-latitude peatlands, which are crucial carbon-sequestering ecosystems at risk to climate change effects. It has been previously suggested that microbial methane-fueled fixation of atmospheric nitrogen (N 2 ) may occur in these ecosystems, but this process and the organisms involved are largely uncharacterized