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Large Scale Oligonucleotide Synthesis

Large scale oligonucleotide synthesis
Large scale oligonucleotide synthesis

Biol-Synthesis offers multi-gram large scale oligonucleotide synthesis, competitive pricing and fast turnaround.

With over 30 years of experience in oligonucleotide services, Bio-Synthesis provides large-scale custom oligonucleotide synthesis for therapeutic projects worldwide. Using various synthesis technologies, we manufacture highly complex DNA and RNA oligonucleotides, which are produced for high-throughput, in vivo or commercial applications in multi-gram quantities of highly purified antisense oligos, siRNAs, ribozymes, miRNA and qPCR primers and probes. All large scale oligonucleotides are manufactured under strict quality management standards, which include MS, HPLC and gel electrophoresis after completing synthesis, purification, and quality control steps. These steps are followed by independent QA (Quality Assurance) procedures which guarantee the highest quality large scale DNA or RNA oligonucletoides.

Large Scale DNA and RNA Oligonucleotide Synthesis

  • mg to 50 gram/batch
  • Single or duplex format
  • Wide range of modifications
  • Contract Research Study
  • Custom analytic method development
  • Custom formulation and packaging

Services

  • We develop custom applications and guide you in transitioning your pilot batch to large-scale production
  • Flexible ordering methods
  • Dedicated purification column
  • Custom aliquoting and vialing formats
  • Product compliance verification and release prior to shipment
  • Custom secure online portal for production status review
  • Digital or hard copy of analytical QC documentation
Backbone Options

We are able to incorporate various types of modifications specifically for antisense or RNAi applications. The most popular modification used for these applications involves replacing Phophodiester with Phosphothiolate linkages. Such linkages are highly resistant to nuclease degradation, but they also lower duplex stability. To compensate for such deficiency, we can incorporate modified bases which do not only minimize exonuclease degradation but also enhance duplex stability. See examples below.

  • Bridged Nucleic Acid (BNA)
  • 2' Fluoro RNA
  • 2'-F-ANA
  • 2' O-Methyl RNA (2'OMe)
  • 2' O-Methyoxy Ethyl
  • Phosphorothioate DNA
  • Phosphorothioate RNA
  • Phosphonoacetate (PACE)
  • 2,6-Diaminopurine-riboside
  • 5-Methylcytidine
  • 2'-Deoxyisoguanosine
  • 2,6-Diaminopurine-2'-O-methylriboside
  • 5-Methyl-2'-O-Methylcytidine
  • 7-Deaza-8-aza-2'-deoxyadenosine
  • 5-Methyl-2'-deoxyisocytidine

 

* We offer custom BNA oligonucleotide synthesis as an alternative to LNA and PNA.

Purification

Several types of purification are available

Standard Purification

Since the salt content and the purity of oligonucleotides are important features during large scale synthesis, a desalting step is needed as an additional default purification method for this service.

HPLC Purification

  • HPLC is well suited for the purification of unmodified or modified oligonucleotides with sequence lengths of up to 60 bases. Therefore HPLC is the method of choice for purifying larger amounts of oligonucleotides (i.e. > 1 mol). A purity of 85% is guaranteed for oligos up to 40 bases.
  • Ion-exchange HPLC (IE-HPLC) is available to purify long oligonucleotides of up to 60 bases. Anion-exchange resins separate oligos on the basis of charge. The resolution of this technique is very high but their use is limited to the purification of shorter oligonucleotides. For unmodified oligos between 11-40 bases, a purity of > 90% is guaranteed.
  • Dual HPLC Purification and Dual PAGE and HPLC Purification can also be applied: Options are the combined use of HPLC and IEX HPLC purification for 20-30 mers oligonucleotides, as well as HPLC and PAGE purification for 31 - 50 mers oligonucleotides. The use of combined purifications enable average purities of 98% allowing the removal of most base defective products and other impurities.
  • RNAase Free HPLC Purification, originally developed for isolating RNA oligos and RNase-Free HPLC Purification, has been extended to DNA oligonucleotides for its use when there is a need for oligonucleotides free of ribonucleases. RNase-Free HPLC purification is carefully performed in an RNase-Free environment; reagents, equipment and lab surfaces are monitored for RNase contamination.

In-Vivo-Ready Purification

Our In-Vivo-Ready purification option provides high quality oligonucleotides that are ready for animal studies. Each oligonucleotide is HPLC purified, dialyzed to ensure that the oligonucleotide is salt free (conductance of <1 microsiemens for a 50 um solution), sterile filtered, and endotoxin tested. All oligonucleotides are analyzed by MALDI-TOF mass spectrometry, analytical HPLC, and sequencing.

Quality Control

Several types of quality control

Standard QC package

  • Analytical anion-exchange HPLC of oligonucleotide; purity guaranteed above 85% in case purification is required.
  • TOF/MS for oligonucleotide identification.

Advanced QC package

  • Standard QC package and additionally:
  • Ion-pair reverse phase HPLC of the oligonucleotide at 60 C, purity guaranteed above 85%
  • Analytical reverse phase HPLC
  • Endotoxin analysis of the oligonucleotide (*phosphorothioates may inhibit in the mechanism for endotoxin determinations and therefore readings might be less accurate for these molecules)

Custom QC Package:

  • Determination of the sodium content of the oligonucleotide (minimum 20 mg of purified oligonucleotide is required).
  • LC-MS of the oligonucleotides
  • Any Custom quality control methods could be developed.
  • Endotoxin analysis of the oligonucleotide (*phosphorothioates may inhibit in the mechanism for endotoxin determinations and therefore readings might be less accurate for these molecules)
Applications for single-stranded and douple-stranded oligonucleotides
  • qPCR Controls
  • Gene Construction or Assembly
  • Use of oligonucleotides for the production of recombinant Antibodies
  • Enzyme Substrates
  • in vitro transcription approaches
  • Gene Assembly or gene construction
  • Affinity capture probes
  • And many others
Manufacturing Line Compliant with GMP

Our quality products are manufactured in compliance with GMP standards.

Pricing

Our pricing applies to unmodified oligonucleotides between 15 and 25 bases. Oricing includes purifications and ethanol precipitation. Please request a quotation for other lengths or larger quantities. The backbone may consist of phosphorothioate or phosphodiester linkages, or a combination of the two. Prices may vary due to sequence composition.