Oligo Backbone Modification

Oligo Backbone Modification

BOC Sciences offers the synthesis of a variety of backbone modified oligonucleotides, to increase the affinity of the oligonucleotide for its complementary strand, which often significantly improves the specificity and sensitivity of applications that detect small or highly similar DNA or RNA targets. These backbone modified oligonucleotides not only provide strong affinity enhancement, but also increase resistance to endonucleases and exonucleases, resulting in high in vitro and in vivo stability.


The oligonucleotide backbone consists of phosphodiester linkages and sugar moieties, which can be properly modified to improve the properties of various oligonucleotides, including improved stability to nuclease and uptake into cells, increased affinity for complementary strands, enhanced kinetics and base pairing specificity and sensitivity to binding to nucleic acid targets.

  • Phosphorothioate (PS) modification

The synthesis of phosphorothioate oligonucleotides is achieved by replacing the non-bridging oxygen in the phosphate backbone of the oligonucleotide with a sulfur atom. PS-linkage modification has two excellent advantages. The first is to increase the stability of nuclease digestion. This modification will prolong the half-life of the oligonucleotide sequence and prevent its degradation. The second is to enhance the binding of proteins, especially serum proteins. Serum interactions include high-affinity interactions with heparin-binding proteins and low-affinity interactions with albumin. Increased binding to serum proteins retains circulating oligonucleotides, slows hepatic clearance, and prolongs absorption into target tissues. These advantages have led to their widespread use in the manufacture of first-generation antisense oligonucleotides (ASOs), which are designed to inhibit the expression of target RNAs through sequence-specific recognition. However, ASOs have the significant disadvantage of being severely toxic and prone to apoptosis.

  • Phosphorodithioate modification

The two non-bridging oxygen atoms of the oligonucleotide are replaced by sulfur atoms. These linkages are achiral and are fully resistant to cleavage by all known nucleases. However, phosphorodithioate-modified oligonucleotides are less used due to their reduced recognition ability when combined with complementary oligonucleotides, and can also bind to various proteins.

  • Methylphosphonate modification

Methylphosphonate oligodeoxynucleotides (MPO), a metabolically stable analogue of conventional DNA, contain a methyl group replacement for a non-bonded phosphoryl oxygenate. This modification imparts a neutral charge to the methyl phosphonate backbone linkage. Oligonucleotides containing one or more methylphosphonate bonds will resist nuclease degradation at these sites, while the lack of charge will improve intracellular transport. Based on these properties, MPO has been explored as antisense reagents.

  • 2'-ribose modification

Ribose can be modified by replacing the 2'-hydroxyl group with different groups, but the most common are 2'-O-methyl, 2'-O-methoxyethyl and 2'-fluorine. The 2' ribose modification enhanced stability to nuclease digestion by blocking the nucleophilic 2' hydroxyl moiety. These modifications also increase the thermal stability of complementary hybridization, promote tighter binding and allow the use of shorter oligonucleotides. The fully modified oligonucleotide at the 2' position does not support RNase H activity and therefore is not cleaved, this property that allows it to be employed to protect mRNA targets and is currently exploited to design oligomers that regulate alternative splicing. The modifications at the 2' position are well tolerated in oligonucleotide duplexes, which enable the 2'-position to bind various functional groups, such as fluorophores.

  • Mesyl phosphoramidate modification

The oligodeoxynucleotide replaces the phosphodiester (PO) group with a methanesulfonyl (methanesulfonyl) phosphoramidate group and exhibits high affinity for RNA, excellent resistance to nucleases, efficient recruitment of RNase H, and higher efficiency in catalyzing the degradation of oncogenic microRNAs (miRNAs). These remarkable features have led to its application in creating improved ASOs with potent anticancer effects.

  • Bridged Nucleic Acids (BNA)

The 2'-oxygen can also bind to the 4'-carbon of the ribose by bridging the carbon to form BNA. The first generation of BNAs are locked nucleic acids (LNAs) characterized by the presence of a covalent bond between the 2' oxygen and 4' carbon in the ribose. BNAs can readily bind to oligonucleotide chains, which can improve hybridization strength. BNA oligonucleotides show better base stacking and thermal stability, leading to extremely efficient binding to complementary nucleic acids and upgraded mismatch recognition. The introduction of a single BNA substitution can increase the melting temperature by 5-10°C, allowing the affinity for complementary hybridization to be tailored for clinical applications. BNA oligonucleotides also exhibit enhanced resistance to nucleases and are easily labeled or modified with standard oligonucleotide tags such as DIG, fluorescent dyes, biotin, amino adapters, etc. BNA oligos have been successfully used for single nucleotide polymorphism (SNP) detection and analysis, primer and probe design, and in antigene technology.

Our Oligo Backbone Modification Services Specification

ModificationsNuclease ResistanceRNase H ActivationPrice
Bridged Nucleic Acids (BNA)Inquiry
2' Fluoro RNAInquiry
2' O-Methyl RNA (2'OMe)Inquiry
Phosphorothioate DNAInquiry
Phosphorothioate RNAInquiry
Phosphonoacetate (PACE)Inquiry
Methylphosphonate linkagesInquiry
ZNA SpermineInquiry

Our oligonucleotide backbone modification synthesis services provide:

  • Scale: 100 nmole, 250 nmole, 1 umole or larger.
  • Purification options: desalting, PAGE, HPLC, dual HPLC
  • Oligonucleotide length: 5-100 bases
  • Delivery method: Tube or plate lyophilized
  • Provides concentration adjustments and additional aliquots
  • By OD measurement and quality control


  • High-quality products
  • Competitive price
  • Data analysis, detailed reporting including results and discussion
  • All samples are carefully monitored for stability and characterized to ensure batch-to-batch consistency


  1. Micklefield, J; et al. Backbone modification of nucleic acids: synthesis, structure and therapeutic applications. Curr Med Chem. 2001. 8(10): p. 1157-79.
  2. Shen, X; et al. Chemistry, mechanism and clinical status of antisense oligonucleotides and duplex RNAs. Nucleic Acids Research. 2017. 46(4): p. 1584-1600.
  3. Patutina, O.A.; et al. Mesyl phosphoramidate backbone modified antisense oligonucleotides targeting miR-21 with enhanced in vivo therapeutic potency. Proc Natl Acad Sci U S A. 2020. 117(51): p. 32370-32379.
* Only for research. Not suitable for any diagnostic or therapeutic use.
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