Our phosphate backbone-modified siRNA services support research teams developing chemically stabilized RNA interference reagents for target validation, functional genomics, delivery studies, and discovery-stage candidate optimization. By replacing or modifying selected phosphodiester linkages, researchers can adjust nuclease resistance, metabolic stability, charge distribution, protein interactions, and strand behavior without changing the underlying siRNA sequence.
We provide sequence- and position-specific backbone design, custom synthesis, purification, duplex preparation, analytical verification, and optional functional testing. Projects may include terminal phosphorothioate protection, mixed phosphodiester/phosphorothioate patterns, phosphorodithioate or boranophosphate feasibility studies, phosphonate-based linkages, and stable 5′-phosphate analog strategies. These options can also be coordinated with ribose-modified siRNA, terminal functionalization, labeling, or conjugation when a multilayered chemistry design is required.
Figure 1. Phosphate backbone modifications of siRNA.
Premature Duplex Degradation: Native phosphodiester linkages can be vulnerable to exonucleases and endonucleases in biological matrices. We evaluate terminal protection, overhang stabilization, and strand-specific linkage placement to improve the usable experimental window without automatically modifying every internucleotide bond.
Loss of RNAi Activity: Backbone changes that are placed too densely or at functionally sensitive positions may interfere with duplex unwinding, Ago2 loading, target recognition, or passenger-strand removal. Our design process considers guide-strand orientation, seed-region sensitivity, terminal recognition, and intended RNAi mechanism before a modification map is finalized.
Unbalanced Strand Protection: The guide and passenger strands do not always require identical modification patterns. We develop strand-resolved architectures that can protect vulnerable termini while preserving the asymmetry needed for productive guide-strand selection and limiting unwanted passenger-strand activity.
Complex Synthesis and Analysis: Sulfur-rich, mixed-linkage, and advanced backbone constructs may display different coupling behavior, purification profiles, stereochemical complexity, and mass-spectrometric characteristics from standard RNA. We align synthesis, purification, and analytical methods with the selected backbone chemistry and modification density.
Interaction With Other Modifications: Backbone chemistry cannot be evaluated independently when the duplex also contains 2′-O-methyl, 2′-fluoro, terminal ligands, hydrophobic groups, fluorophores, or delivery components. Our integrated planning connects backbone placement with chemically modified siRNA and siRNA conjugate workflows to reduce avoidable compatibility problems.
Our services cover the complete development path from backbone strategy and candidate-panel planning to synthesis, duplex assembly, analytical characterization, and research-use testing. Each project is reviewed according to the target sequence, guide and passenger strand architecture, degradation pressure, delivery format, and required experimental readout.
Rather than applying a standard modification pattern to every sequence, we support position-specific designs and comparative panels that help researchers determine how linkage type, density, and strand distribution affect stability and RNAi performance.
| Modification | Quantity | Purification | Details | Price |
| 5'-Bromo-rU | 2 OD-250 OD | HPLC | A bromine atom is introduced at the uridine position at the 5' end of the RNA molecule, which is mainly used for cellular and tissue tracing. | Inquiry |
| 5'-Phosphate | 2 OD-250 OD | HPLC | The introduction of a phosphate group at the 5' end backbone of siRNA enables regulation of stability and affinity and further regulation of transcription and translation. | Inquiry |
| 5'-Amine | 2 OD-250 OD | HPLC | Modification by introducing an amine (NH2) group at the 5' end backbone of siRNA may be useful for subsequent bioconjugation. | Inquiry |
| 5'-Biotin | 2 OD-250 OD | HPLC | The modification of introducing biotin molecules into the 5' end backbone of siRNA can be used to track and localize the effects of RNAi. | Inquiry |
| 5'-Thiol | 2 OD-250 OD | HPLC | This modification increases the half-life of siRNA molecules in biological systems, resulting in more effective and sustained gene silencing. | Inquiry |
| 5'-Ferrocene | 2 OD-250 OD | HPLC | Attachment of a ferrocene group to the 5' end of an siRNA molecule enhances stability and is widely used for bioimaging, electrochemical detection, cell tracing and gene therapy. | Inquiry |
| 5'-Phosphorothioate | 2 OD-250 OD | HPLC | The replacement of one or more non-bridging oxygen atoms in the phosphate backbone of siRNA by sulfur atoms enhances the stability and extends the half-life of siRNA. | Inquiry |
| 5'-Phosphorodithioate | 2 OD-250 OD | HPLC | Phosphomethylation modification can increase the stability and intracellular stability of siRNA as well as improve the targeting effect. | Inquiry |
| 5'-Phosphoramidate | 2 OD-250 OD | HPLC | The introduction of alcohol ester group modification on the phosphate group in the phosphate backbone improves the stability of siRNA. | Inquiry |
| 5'-Phosphorothioate methyl ester | 2 OD-250 OD | HPLC | The introduction of methyl ester group on the sulfur atom of the phosphate backbone of siRNA can change the hydrophilicity and lipid solubility of siRNA and increase the intracellular stability. | Inquiry |
| 5'-Phosphocholine | 2 OD-250 OD | HPLC | The introduction of choline groups onto the phosphate backbone improves the stability and cell permeability of siRNAs and helps to enhance the biodistribution and transport of siRNAs. | Inquiry |
| 5'-Phosphoglycerol | 2 OD-250 OD | HPLC | Phosphoglycerol modification can alter the water solubility, stability and cell permeability of siRNA. | Inquiry |
| 5'-Phosphoethoxy | 2 OD-250 OD | HPLC | Replacement of one or more phosphate groups on the phosphate backbone using vinyl groups can alter the physical and chemical properties, stability and affinity of siRNA. | Inquiry |
Backbone chemistries differ in charge, stereochemical behavior, nuclease resistance, Ago2 compatibility, and synthesis complexity. The appropriate choice depends on whether the project is focused on terminal protection, mixed-linkage optimization, mechanistic research, or comparison of emerging backbone concepts.
| Backbone Option | Structural Feature | Primary Design Use | Key Considerations | Suitable Project Types |
| Phosphodiester (PO) | Native negatively charged internucleotide linkage | Preserve natural RNA geometry and provide a reference for modification studies | More vulnerable to nuclease cleavage; often retained at functionally sensitive internal positions | Control duplexes, baseline potency studies, mixed-backbone designs |
| Phosphorothioate (PS) | One non-bridging phosphate oxygen is replaced with sulfur | Protect termini or selected linkage regions from degradation | Introduces phosphorus stereochemistry; placement and density may affect protein interactions and RNAi activity | Terminal protection, overhang stabilization, chemically stabilized duplexes |
| Phosphorodithioate (PS2) | Both non-bridging phosphate oxygens are replaced with sulfur | Explore sulfur-rich linkages with reduced phosphorus stereocomplexity | Effects are position-dependent; synthesis, purification, and analytical methods require dedicated planning | Advanced backbone screening, terminal-linkage comparisons, chemistry research |
| Boranophosphate | A non-bridging phosphate oxygen is replaced with a borane group | Investigate an alternative nuclease-resistant backbone while retaining an anionic linkage | Availability, placement tolerance, and functional performance must be assessed sequence by sequence | Mechanistic studies, advanced candidate panels, comparative stability research |
| Alkyl Phosphonate | A non-bridging oxygen is replaced with an alkyl substituent, reducing local charge | Tune local backbone charge or investigate position-specific specificity effects | Internal placement may alter duplex structure, Ago2 interactions, solubility, or target recognition | Seed-region research, off-target mechanism studies, structure-activity screening |
| 5′-Phosphate Mimic | Metabolically stabilized analog of the guide-strand 5′ phosphate | Preserve an Ago2-recognized terminal feature under degradation pressure | Terminal geometry and electronic properties must remain compatible with guide-strand loading | Guide-strand optimization, terminal stability studies, advanced siRNA architectures |
The same backbone linkage can produce different outcomes depending on its location. This matrix summarizes common placement objectives and the issues that should be considered before guide- and passenger-strand modification patterns are finalized.
| Modification Region | Typical Objective | Primary Design Risk | Recommended Evaluation | Project Deliverable |
| Guide 5′ Terminus | Protect a functionally important terminus while maintaining productive Ago2 recognition | Poorly selected linkages or terminal groups may reduce guide loading | Terminal-chemistry review, matched variant comparison, RNAi activity testing | Annotated guide-strand terminal design |
| Guide 3′ Terminus | Reduce degradation while preserving PAZ-domain-compatible strand behavior | Excessive modification may change end recognition or duplex release | Stability testing, duplex analysis, functional comparison | Guide 3′-end linkage map |
| Passenger 5′ Terminus | Control passenger-strand stability and influence strand-selection behavior | Unintended stabilization may increase passenger-strand participation | Thermodynamic asymmetry review and strand-specific activity assessment | Passenger-strand protection strategy |
| Passenger 3′ Terminus | Improve duplex persistence and protect exposed terminal linkages | An overly stable passenger strand may hinder duplex unwinding | Annealing review, degradation study, RNAi comparison | Passenger 3′-end linkage map |
| 3′ Overhangs | Protect exposed nucleotides commonly susceptible to exonuclease attack | Overhang chemistry may affect terminal recognition and duplex handling | Overhang-format comparison and matrix stability testing | Overhang modification specification |
| Guide Seed Region | Investigate position-specific effects on specificity and unintended transcript recognition | Internal backbone changes may reduce on-target cleavage or distort Ago2 interactions | Small candidate panel with on-target and specificity-focused testing | Seed-region structure-activity panel |
| Conjugation Junction | Protect linkages adjacent to a ligand, linker, fluorophore, lipid, or peptide attachment | Combined hydrophobicity, steric effects, and backbone chemistry may complicate purification or activity | Conjugate-aware design, analytical review, and matched unconjugated control | Integrated conjugate and linkage map |
| Internal Duplex Core | Increase broader resistance or study novel backbone effects | Dense internal modification may interfere with duplex geometry, cleavage, or RISC processing | Limited initial placement, structural review, and empirical activity screening | Staged internal-modification plan |
Our workflow connects siRNA sequence design with realistic chemistry, analytical, and functional requirements. Each stage is intended to reduce ambiguity around strand identity, linkage placement, candidate comparisons, and final research deliverables.
We confirm the target sequence, intended experimental system, guide and passenger strands, preferred duplex architecture, degradation challenge, delivery format, required quantity, and downstream assays. Existing sequence or potency data are reviewed when available.
The sequence is evaluated for terminal exposure, guide-strand 5′-end requirements, overhang design, modification-sensitive regions, and compatibility with any ribose, base, label, linker, or conjugate components. Potential synthesis and purification risks are identified before the design is locked.
We prepare a strand-resolved modification map and recommend either a single construct or a comparative panel. The proposal defines backbone type, linkage positions, control candidates, purification approach, analytical methods, and optional stability or RNAi testing.
Guide and passenger strands are synthesized using conditions appropriate for their backbone chemistry and modification density. Purification is selected to separate truncated sequences, incompletely modified material, and other process-related impurities relevant to the construct.
Purified strands are characterized and annealed in the required ratio and format. Identity, purity, concentration, strand documentation, and agreed duplex-related attributes are reviewed before the material proceeds to optional stability or functional studies.
The final package includes sequence information, guide/passenger annotation, linkage maps, analytical results, handling details, and optional comparative testing data. Follow-up support can address candidate ranking, expanded backbone panels, conjugation, or additional siRNA screening.
Backbone engineering requires more than adding sulfur linkages to a sequence. Our platform combines siRNA mechanism awareness, modification mapping, synthesis planning, analytical verification, and optional functional comparison so that each construct can be evaluated as a complete research reagent.
Phosphate backbone-modified siRNA can support research programs that require greater resistance to degradation, controlled strand behavior, advanced chemical architecture, or direct comparison of linkage strategies. Modification patterns should be selected according to the experimental system and validated rather than assumed to improve every sequence.
Whether your project requires terminal phosphorothioate protection, an asymmetric mixed-backbone duplex, a comparative modification panel, an advanced phosphate mimic, or an integrated backbone-and-ribose design, our team can help translate the concept into a clearly specified research reagent. We support sequence review, modification mapping, custom synthesis, purification, duplex assembly, analytical verification, stability testing, and optional RNAi evaluation. Contact us with your target sequence, preferred duplex format, modification goals, quantity requirements, and planned assays to begin a technical assessment.
Phosphate backbone modifications significantly enhance nuclease resistance, improve serum stability, extend half-life in biological systems, and maintain effective gene silencing activity.
We provide comprehensive modifications including phosphorothioate, phosphorodithioate, phosphoramidate, and specialized groups like biotin, thiol, and ferrocene for various research applications.
Strategic backbone modifications improve membrane permeability, enhance biodistribution, and facilitate intracellular delivery while maintaining RNA interference efficiency.
Yes, we specialize in creating hybrid modification strategies that integrate backbone modifications with base and ribose modifications for optimal siRNA performance and stability.
All backbone-modified siRNAs undergo rigorous HPLC purification, mass spectrometry verification, and functional validation to guarantee purity, stability, and biological activity.
We provide lyophilized powders in various quantities (2 OD-250 OD) with flexible packaging options, including formats ready for direct research applications.
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