Fully Automated Parallel Oligonucleotide Synthesizer

2001 ◽  
Vol 66 (8) ◽  
pp. 1299-1314 ◽  
Author(s):  
Michal Lebl ◽  
Christine Burger ◽  
Brett Ellman ◽  
David Heiner ◽  
Georges Ibrahim ◽  
...  
Keyword(s):  
Solid Phase ◽  
Building Blocks ◽  
Solid Support ◽  
Continuous Operation ◽  
Gear Pump ◽  
Oligonucleotide Synthesis ◽  
Center Of Rotation ◽  
Microtiter Plates ◽  
The Individual ◽  
Design And Construction

Design and construction of automated synthesizers using the tilted plate centrifugation technology is described. Wash solutions and reagents common to all synthesized species are delivered automatically through a 96-channel distributor connected to a gear pump through two four-port selector valves. Building blocks and other specific reagents are delivered automatically through banks of solenoid valves, positioned over the individual wells of the microtiterplate. These instruments have the following capabilities: Parallel solid-phase oligonucleotide synthesis in the wells of polypropylene microtiter plates, which are slightly tilted down towards the center of rotation, thus generating a pocket in each well, in which the solid support is collected during centrifugation, while the liquid is expelled from the wells. Eight microtiterplates are processed simultaneously, providing thus a synthesizer with a capacity of 768 parallel syntheses. The instruments are capable of unattended continuous operation, providing thus a capacity of over two millions 20-mer oligonucleotides in a year.

New cleavable spacers for tandem synthesis of multiple oligonucleotides

Synthesis ◽  
2021 ◽  
Author(s):  
Kazuki Yamamoto ◽  
Yasufumi Fuchi ◽  
Masaya Okabe ◽  
Takashi Osawa ◽  
Yuta Ito ◽  
...  
Keyword(s):  
Aqueous Ammonia ◽  
Solid Phase ◽  
Solid Support ◽  
Oligonucleotide Synthesis ◽  
Single Column ◽  
Tandem Synthesis ◽  
Single Sequence

In solid-phase oligonucleotide synthesis, a single oligonucleotide is generally acquired from a column loaded with a specific solid support. Herein, we have developed new cleavable spacer (CS) derivatives for tandem synthesis of multiple oligonucleotides on a single column. Four CS analogs were designed, synthesized, and inserted between two oligonucleotide sequences using an automated oligonucleotide synthesizer. The CS derivatives bearing a cyclic cis-1,2-diol exhibited efficient release of the two oligonucleotides under commonly employed basic conditions of aqueous ammonia. Among the CS analogues, it was found that CS with a robust structure can potentially be applied as a spacer molecule in the tandem synthesis of multiple oligonucleotides in a single sequence.

3-(Tributylstannyl)allyl Alcohols: Useful Building Blocks for Solid-Phase Synthesis of Skipped Dienes and Trienes

2000 ◽  
Vol 65 (3) ◽  
pp. 434-454 ◽  
Author(s):  
Miroslav Havránek ◽  
Dalimil Dvořák
Keyword(s):  
Solid Phase Synthesis ◽  
Solid Phase ◽  
Building Blocks ◽  
Solid Support ◽  
Stille Coupling ◽  
Phase Synthesis ◽  
Negative Results ◽  
Allyl Alcohols

Repeated Stille coupling of 3-substituted 3-(tributylstannyl)allyl alcohols 2 on a solid support was used to synthesize a 21 × 21 library of skipped dienes and a 21 × 21 × 21 library of skipped trienes. Starting 3-(tributylstannyl)allyl alcohols were prepared by Pd-catalyzed hydrostannation of substituted prop-2-yn-1-ols, by hydroalumination by LiAlH4 followed with transmetallation to tin using tributyltin methoxide, or by substitution of chlorine in (Z)-6-chloro-3-(tributylstannyl)hex-2-en-1-ol with appropriate nucleophile. Synthesized libraries were tested for the activity to endorphin receptors, but with negative results.

Scalable One-Pot - Liquid-Phase Oligonucleotide Synthesis for Model Network Hydrogels

2020 ◽  
Author(s):  
Guido Creusen ◽  
Cecilia Oluwadunsin Akintayo ◽  
Katja Schumann ◽  
Andreas Walther
Keyword(s):  
Liquid Phase ◽  
Solid Phase ◽  
Adenine Nucleotides ◽  
Building Blocks ◽  
Oligonucleotide Synthesis ◽  
One Pot ◽  
Model Network ◽  
Dna And Rna ◽  
Sequence Recognition ◽  
Sequential Coupling

Solid-phase oligonucleotide synthesis (SPOS) based on phosphoramidite chemistry is currently the most widespread technique for DNA and RNA synthesis, but suffers from scalability limitations and high reagent consumption. Liquid-phase oligonucleotide synthesis (LPOS) uses soluble polymer supports and has the potential of being scalable. However, at present, LPOS requires 3 separate reaction steps and 4-5 precipitation steps per nucleotide addition. Moreover, long acid exposure times during the deprotection step degrade sequences with high A-content (adenine) due to depurination and chain cleavage. In this work, we present the first one-pot liquid-phase DNA synthesis technique, which allows the addition of one nucleotide in a one-pot reaction of sequential coupling, oxidation and deprotection, followed by a single precipitation step. Furthermore, we demonstrate how to suppress depurination during the addition of adenine nucleotides. We showcase the potential of this technique to prepare high-purity 4-arm PEG‑T<sub>20</sub> (T = thymine) and 4-arm PEG-A<sub>20</sub>building blocks in multi-gram scale. Such complementary 4-arm PEG-DNA building blocks reversibly self-assemble into supramolecular model network hydrogels, and facilitate the elucidation of bond lifetimes. These model network hydrogels exhibit new levels of mechanical properties, high stability at room temperature (melting at 44 ‎°C), and thus open up pathways to next-generation, scalable DNA-materials programmable through sequence recognition and available for macroscale applications<i>.</i>

scholarly journals Solid Phase Assembly of Fully Protected Trinucleotide Building Blocks for Codon-Based Gene Synthesis

2019 ◽  
Vol 9 (11) ◽  
pp. 2199 ◽  
Author(s):  
Ruth Suchsland ◽  
Bettina Appel ◽  
Matthäus Janczyk ◽  
Sabine Müller
Keyword(s):  
Amino Acids ◽  
Solid Phase ◽  
Building Blocks ◽  
Disulfide Bridge ◽  
Solid Support ◽  
Gene Synthesis ◽  
Directed Mutagenesis ◽  
Reducing Conditions

The use of pre-formed trinucleotides, representing codons of the 20 canonical amino acids, for oligonucleotide-directed mutagenesis offers the advantage of controlled randomization and generation of “smart libraries”. We here present a method for the preparation of fully protected trinucleotides on solid phase. The key issue of our strategy is the linkage of the starting nucleoside to the solid support via a traceless disulfide linker. Upon trinucleotide assembly, the disulfide bridge is cleaved under reducing conditions, and the fully protected trinucleotide is released with a terminal 3′-OH group.

scholarly journals Convergent Synthesis of Thioether Containing Peptides

Molecules ◽  
2020 ◽  
Vol 25 (1) ◽  
pp. 218
Author(s):  
Spyridon Mourtas ◽  
Christina Katakalou ◽  
Dimitrios Gatos ◽  
Kleomenis Barlos
Keyword(s):  
Carboxylic Acids ◽  
Peptide Synthesis ◽  
Solid Phase ◽  
Solid Phase Peptide Synthesis ◽  
Building Blocks ◽  
Solid Support ◽  
Convergent Synthesis ◽  
Fmoc Group ◽  
Synthetic Routes ◽  
Peptide Elongation

Thioether containing peptides were obtained following three synthetic routes. In route A, halo acids esterified on 2-chlorotrityl(Cltr) resin were reacted with N-fluorenylmethoxycarbonyl (Fmoc) aminothiols. These were either cleaved from the resin to the corresponding (Fmoc-aminothiol)carboxylic acids, which were used as key building blocks in solid phase peptide synthesis (SPPS), or the N-Fmoc group was deprotected and peptide chains were elongated by standard SPPS. The obtained N-Fmoc protected thioether containing peptides were then condensed either in solution, or on solid support, with the appropriate amino components of peptides. In route B, the thioether containing peptides were obtained by the reaction of N-Fmoc aminothiols with bromoacetylated peptides, which were synthesized on Cltr-resin, followed by removal of the N-Fmoc group and subsequent peptide elongation by standard SPPS. In route C, the thioether containing peptides were obtained by the condensation of a haloacylated peptide synthesized on Cltr-resin and a thiol-peptide synthesized either on 4-methoxytrityl(Mmt) or trityl(Trt) resin.

Automated glycopeptide assembly by combined solid-phase peptide and oligosaccharide synthesis

2014 ◽  
Vol 50 (15) ◽  
pp. 1851-1853 ◽  
Author(s):  
Mattan Hurevich ◽  
Peter H. Seeberger
Keyword(s):  
Amino Acid ◽  
Solid Phase ◽  
Building Blocks ◽  
Solid Support ◽  
Automated Synthesis ◽  
Oligosaccharide Synthesis ◽  
Polystyrene Beads ◽  
Single Instrument

Automated synthesis of glycopeptides was achieved using monosaccharide and amino acid building blocks. Using polystyrene beads equipped with photo-labile linker as solid support, all synthetic manipulations were performed using a single instrument.

scholarly journals Scalable One-Pot - Liquid-Phase Oligonucleotide Synthesis for Model Network Hydrogels

2020 ◽  
Author(s):  
Guido Creusen ◽  
Cecilia Oluwadunsin Akintayo ◽  
Katja Schumann ◽  
Andreas Walther
Keyword(s):  
Liquid Phase ◽  
Solid Phase ◽  
Adenine Nucleotides ◽  
Building Blocks ◽  
Oligonucleotide Synthesis ◽  
One Pot ◽  
Model Network ◽  
Dna And Rna ◽  
Sequence Recognition ◽  
Sequential Coupling

Solid-phase oligonucleotide synthesis (SPOS) based on phosphoramidite chemistry is currently the most widespread technique for DNA and RNA synthesis, but suffers from scalability limitations and high reagent consumption. Liquid-phase oligonucleotide synthesis (LPOS) uses soluble polymer supports and has the potential of being scalable. However, at present, LPOS requires 3 separate reaction steps and 4-5 precipitation steps per nucleotide addition. Moreover, long acid exposure times during the deprotection step degrade sequences with high A-content (adenine) due to depurination and chain cleavage. In this work, we present the first one-pot liquid-phase DNA synthesis technique, which allows the addition of one nucleotide in a one-pot reaction of sequential coupling, oxidation and deprotection, followed by a single precipitation step. Furthermore, we demonstrate how to suppress depurination during the addition of adenine nucleotides. We showcase the potential of this technique to prepare high-purity 4-arm PEG‑T<sub>20</sub> (T = thymine) and 4-arm PEG-A<sub>20</sub>building blocks in multi-gram scale. Such complementary 4-arm PEG-DNA building blocks reversibly self-assemble into supramolecular model network hydrogels, and facilitate the elucidation of bond lifetimes. These model network hydrogels exhibit new levels of mechanical properties, high stability at room temperature (melting at 44 ‎°C), and thus open up pathways to next-generation, scalable DNA-materials programmable through sequence recognition and available for macroscale applications<i>.</i>

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