Direct oligonucleotide synthesis onto super-paramagnetic beads
| Autor: | Michael S. Akhras, Marilyn Fukushima, Nader Pourmand, Michael A. Jensen, Ronald W. Davis |
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| Rok vydání: | 2013 |
| Předmět: |
Technology
Oligonucleotides Bioengineering Chemistry Techniques Synthetic Oligonucleotide synthesis Applied Microbiology and Biotechnology Article Dynabeads chemistry.chemical_compound Automation Synthesis automation Engineering Genetics Peptide bond Chromatography High Pressure Liquid Phosphoramidite Chromatography Peptide nucleic acid DNA synthesis Chemistry Oligonucleotide Synthetic General Medicine DNA Chemistry Techniques Biological Sciences Combinatorial chemistry Magnetic Fields Nucleic acid High Pressure Liquid Super-paramagnetic beads Indicators and Reagents Biotechnology |
| Zdroj: | Journal of biotechnology, vol 167, iss 4 |
| Popis: | Super-paramagnetic beads (SPMB)s used for a variety of molecular diagnostic assays are prepared by attaching pre-synthesized oligonucleotides to the surface via a cumbersome and low efficient method of carbodiimide-mediated amide bond formation. To mainstream the process, we describe a novel procedure of direct oligonucleotide synthesis onto the surface of SPMBs (e.g. MyOne Dynabeads). With the many challenges surrounding containment of paramagnetic beads (≤ 1 μm) during automated oligonucleotide synthesis, we show that by applying a magnetic force directly to the SPMBs we prevent their loss caused by high-pressure drain steps during synthesis. To date we have synthesized 40mers using a Spacer 9 phosphoramidite (triethylene glycol) coupled to the surface of hydroxylated SPMBs. HPLC analysis shows successful product generation with an average yield of 200 pmoles per sample. Furthermore, because of the versatility of this powerful research tool, we envision its use in any laboratory working with conventional synthesis automation, as employed for single columns and for multi-well titer plates. In addition to direct synthesis of oligodeoxynucleotides (DNA) onto SPMBs, this platform also has the potential for RNA and peptide nucleic acid synthesis. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
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