ODN 2006-G5

Class B CpG oligonucleotide - Human TLR9 ligand

ODN 2006-G5 is a B-class CpG ODN with a preference for human TLR9.

CpG ODNs are synthetic oligonucleotides that contain unmethylated CpG dinucleotides in particular sequence contexts (CpG motifs).

These CpG motifs are present at a 20-fold greater frequency in bacterial DNA compared to   mammalian DNA.

CpG ODNs are recognized by Toll-like receptor 9 (TLR9)   leading to strong immunostimulatory effects.

Specifications

Specificity: human TLR9 agonist

Working concentration: 5 µM (10 μg/ml)

Solubility:  5 mg/ml in water

ODN 2006-G5 sequence : 5’-TCGTCGTTTTGTCGTTTTGTCGTTGGGGG-3’ (29 mer)
Note: All bases are phosphodiester.

Quality control:

• TLR9 activity has been tested using HEK-Blue™ TLR9 cells.
• The absence of bacterial contamination (e.g. lipoproteins and endotoxins) has been confirmed using HEK-Blue™ TLR2 and HEK-Blue™ TLR4 cells.

Contents

ODN 2006-G5 is provided lyophilized and is available in three quantities.

tlrl-2006g5 (formerly tlrl-hodnbg):

• 200 μg (21.22 nmol) lyophilized ODN 2006-G5
• 1.5 ml sterile endotoxin-free water

tlrl-2006g5-1 (formerly tlrl-hodnbg-1):

• 1 mg (106.1 nmol) lyophilized ODN 2006-G5
• 1.5 ml sterile endotoxin-free water

tlrl-2006g5-5 (formerly tlrl-hodnbg-5):

• 5 mg (530.5 nmol) lyophilized ODN 2006-G5
• 10 ml sterile endotoxin-free water

ODN 2006-G5 is shipped at room temperature.

Upon receipt, store at -20 °C.

Description

Bacterial and viral DNA induce strong immunostimulatory effects through the activation of TLR9 due to the presence of unmethylated CpG dinucleotides in particular sequence contexts [1,2]. TLR9 activation can be mimicked by synthetic CpG oligodeoxynucleotides (ODNs). Two types of immunostimulatory CpG-ODNs have been described. Type A (or D) ODNs preferentially activate plasmacytoid dendritic cells (pDC) to produce IFNa, whereas type B (or K) ODNs induce the proliferation of B cells and the secretion of IgM and IL-6 [3]. Another type has been generated that combines features of both types A and B termed type C [4].
Type A ODNs are characterized by a central CpG-containing palindromic motif and a 3’ poly-G string that further increases their activity. They are chimeric ODNs with a central phosphodiester (PD) backbone and flanking phosphorothiate (PTO) modifications. Type B ODNs contain multiple CpGs, preferentially located at the 5’ end, and a PTO backbone.
Cellular uptake is a prerequisite for CpG-induced signal transduction as TLR9 is expressed in the endosome. Uptake of CpG-ODNs in mice is independent of CpG motifs while signaling is strictly dependent on such structures. Indeed, PTO ODNs are taken up much more efficiently than their PD counterparts. However, PTO ODNs are associated with CpG-independent immuno-stimulatory effects and seem to induce a slightly different profile than PD ODNs. Type B prototype ODN 2006 in its PD form is poorly internalized. Addition of a 3’ poly-G string (ODN 2006-G5) was reported to improve its internalization which was correlated with increased IL-6 secretion and PBMC proliferation [5]. Thus, internalization of PD ODNs can be improved by adding a 3’ poly-G string with no need for PTO modifications.

1. Krieg, A.M. et al., 1995. CpG motifs in bacterial DNA trigger direct B-cell activation. Nature, 374(6522):546-9.
2. Bauer, S. et al., 2001. Human TLR9 confers responsiveness to bacterial DNA via species-specific CpG motif recognition. Proc Natl Acad Sci U S A, 98(16):9237-42.
3. Krug A. et al., 2001. Identification of CpG oligonucleotide sequences with high induction of IFN-alpha/beta in plasmacytoid dendritic cells. Eur J Immunol, 31(7): 2154-63.
4. Marshall JD. et al., 2005. Superior activity of the type C class of ISS in vitro and in vivo across multiple species. DNA Cell Biol. 24(2):63-72.
5. Bartz H. et al., 2004. Poly-guanosine strings improve cellular uptake and stimulatory activity of phosphodiester CpG oligonucleotides in human leukocytes.Vaccine. 23(2):148-55.