Oligo Modifications
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Synthetic CpG ODNs activate immune cells through the Toll-like receptor (TLR) pathway

Obtain CpG ODN 1826, CpG ODN 2006, and others with modified backbones

Quick facts:

Ordering path: Order as Custom DNA Oligo

Length: 18–30 bases

Modification: Phosphorothioate bonds

Scales: 100 nmol to large scale synthesis

Purification: Standard desalt; HPLC available upon request

Optional services: Custom preparative or analytical services available upon request, such as Na+ salt exchange, analytical RP-HPLC, endotoxin analysis, aliquot service, etc.

Enhancing the immune response

Originally described in the 1990s, synthetic CpG oligodeoxynucleotides (ODNs) have been extensively studied as an adjuvant to enhance the immune response of vaccines [1]. By mimicking the immune-stimulatory effects of unmethylated bacterial or viral sequences, these synthetic oligos are potent in activating pattern recognition receptors (PRR), promoting cytokine secretion, and, as a result, mounting rapid responses to microbial pathogens.

Structure specific sequences correspond with distinct actions

So far 3 major classes of CpG ODNs have been identified, based on their structural and biological characteristics, and are designated Class A, Class B, and Class C (Table 1) [2]. Class A oligos, which feature a central palindromic CpG-containing phosphodiester (PO) structure followed by a phosphorothioate (PS) homopolymeric G-stretch, are robust inducers of interferon-α (IFN-α) production and dendritic cell maturation [3]. Class B oligos, in contrast, usually contain a full phosphorothioate (PS) backbone. These oligos also stimulate IFN-α production, but to a lesser extent. However, they strongly activate B cells [4]. Class C oligos combine the properties of Class A and B, and are characterized by their complete PS backbone and palindromic CpG-containing motifs. All the CpG ODNs contain one or more unmethylated CpG dinucleotides in specific sequence contexts, which are readily recognized by mammalian cells as an indication of microbial invasion, due to the rarity of this structure in mammalian genomes [5].


Class Structure Stimulation
B cells Plasmacytoid dendritic cells IFN-α
A Central palindromic PO backbone with PS ends Weak Strong Strong
B Linear, full PS Strong Strong Weak
C Palindromic motif, full PS Strong Intermediate Strong

Table 1. Structural and biological characteristics of the 3 major classes of CpG oligodeoxynucleotides (ODNs). PO = phosphodiester; PS = phosphorothioate.


Immunotherapeutics for cancer

In addition to their roles in augmenting protective effects against infectious diseases, CpG ODNs have demonstrated potential in a wide variety of animal models as a novel immunotherapeutic reagent for the treatment of cancer [6]. Cancer immunotherapy has made tremendous progress in the past decade and numerous clinical trials are currently underway in various phases. Either used alone, or in combination with other therapeutic regimens, CpG ODNs mimic the natural Toll-like receptor (TLR) 9 ligand and provoke the production of a whole array of signaling factors in a coordinated manner. These signaling events ultimately trigger the cascade of immune responses against cancer cells [7].

Species specificity

Interestingly, the CpG motifs appear to be species-specific. For example, the optimal mouse CpG motif is GACGTT, while that for use in human contexts is GTCGTT [8]. One of the frequently requested Class B CpG ODNs, CpG ODN 1826, is a well-defined murine TLR 9 agonist, and is thus widely used in rodent models [9]. This oligo is effective in eliciting mouse B cell proliferation, maturation of antigen-presenting cells, and a polarized Th1-type cell response [10]. CpG ODN 1826 contains 2 CpG dinucleotides, both flanked by -GA at the 5' end and -TT at the 3' end. Its backbone is fully phosphothioated, providing nuclease resistance, as opposed to the natural PO backbone found in bacterial or viral genomes.

Simple to order

Order these CpG ODNs directly through the IDT website as Custom DNA oligos, by simply providing their nucleotide sequences. Use an asterisk (*) notation inserted between bases within your sequence to denote the PS chemical modification. As examples, the aforementioned mouse stimulator CpG ODN 1826 and its human counterpart, CpG ODN 2006, can be ordered from the Custom DNA Oligos ordering page by providing the sequences as shown in Table 2.


Oligo name Species IDT ordering format (5' → 3')
CpG ODN 1826 Mouse T*C*C*A*T*G*A*C*G*T*T*C*C*T*G*A*C*G*T*T
CpG ODN 2006 Human T*C*G*T*C*G*T*T*T*T*G*T*C*G*T*T*T*T*G*T*C*G*T*T

Table 2: Ordering format for 2 popular CpG oligodeoxynucleotides (ODNs) containing phosphorothioate (PS) bond modifications.


If you require any custom preparative or analytical service with the ODNs to be used in your experiment, such as endotoxin analysis or custom aliquot service, please contact us at custcare@idtdna.com. For clinical applications, IDT offers GMP manufacturing services. Learn more.

You can also contact our scientific applications experts at applicationsupport@idtdna.com with questions regarding ODN use or to discuss your experimental design.

References

  1. Coffman RL, Sher A, Seder RA. (2010) Vaccine adjuvants: putting innate immunity to work. Immunity, 33(4):492–503.
  2. Vollmer J, Krieg AM. (2009) Immunotherapeutic applications of CpG oligodeoxynucleotide TLR9 agonists. Adv Drug Deliv Rev, 61(3):195–204.
  3. Krug A, Rothenfusser S, et al. (2001) Identification of CpG oligonucleotide sequences with high induction of IFN-alpha/beta in plasmacytoid dendritic cells. Eur J Immunol, 31(7):2154–2163.
  4. Krieg AM, Yi AK, et al. (1995) CpG motifs in bacterial DNA trigger direct B-cell activation. Nature, 374(6522):546–549.
  5. Hemmi H, Takeuchi O, et al. (2000) A Toll-like receptor recognizes bacterial DNA. Nature, 408(6813):740–745.
  6. Krieg AM. (2007) Development of TLR9 agonists for cancer therapy. J Clin Invest, 2007. 117(5):1184–1194.
  7. Krieg AM. (2008) Toll-like receptor 9 (TLR9) agonists in the treatment of cancer. Oncogene, 27(2):161–167.
  8. Bauer S, Kirschning CJ, et al. (2001) Human TLR9 confers responsiveness to bacterial DNA via species-specific CpG motif recognition. Proc Natl Acad Sci USA, 98(16):9237–9242.
  9. McCluskie MJ, Davis HL. (1999) CpG DNA as mucosal adjuvant. Vaccine, 18(3–4):231–237.
  10. Chu RS, Targoni OS, et al. (1997) CpG oligodeoxynucleotides act as adjuvants that switch on T helper 1 (Th1) immunity. J Exp Med, 186(10):1623–1631.

Product focus—oligos and modifications

Custom oligonucleotides and primers

You can order up to 1 µmol desalted, custom synthesized DNA oligonucleotides and they will be shipped to you the next business day (larger scales are shipped within 5 business days). You can also specify whether to receive them dried down or hydrated, and whether you want them already annealed. Every IDT oligonucleotide you order is deprotected and desalted to remove small molecule impurities. Your oligos are quantified twice by UV spectrophotometry to provide an accurate measure of yield. Standard oligos are also assessed by mass spectrometry for quality you can count on.

Learn more or order now.


Oligo modifications

Review a list of the common modifications IDT can add to oligonucleotides here. Not finding a modification you need on the IDT website? IDT will consider any modification you need. Just send your request to noncat@idtdna.com.


Additional reading

Tips for resuspending and diluting your oligonucleotides—You have received your custom oligonucleotides, and now it is time to resuspend and dilute them. Here are a few tips from our scientists that will help facilitate use of the oligos in your experiments.

Oligonucleotide modifications: Choosing the right mod for your needs—Learn about our broad family of oligonucleotide modifications, and get suggestions for selecting modifications that can help you in your research.

Oligonucleotide modifications that block nuclease degradation—Modification Highlight: Are you working with your oligos in cell culture or in vivo? Find out which modifications can be added to an oligo to limit nuclease degradation.

Which type of oligo purification should I choose?—Do your oligos need purification for your intended application? Use these recommendations based on oligo length, application, yield required, and presence of modifications to determine which oligonucleotide purification method to select.

Need a non-standard modification?—Need a modification you don’t find on our website? IDT offers 89 modifications that are not listed in our online catalog. A few of the more popular ones are described along with information on how to order them. IDT will consider any modification you have in mind. Just make a request at noncat@idtdna.com.


Review other DECODED Online newsletter articles on oligo handling and analysis, and oligo modifications.


You can also browse our DECODED Online newsletter for additional application reviews, lab tips, and citation summaries to facilitate your research.


Author: Brian Wang, PhD, is the Genomic Tools Market Development Manager at IDT.

© 2017 Integrated DNA Technologies. All rights reserved. Trademarks contained herein are the property of Integrated DNA Technologies, Inc. or their respective owners. For specific trademark and licensing information, see www.idtdna.com/trademarks.


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