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Custom siRNA Synthesis and Modification
We provide custom siRNA synthesis with chemical modifications and fluorescent labels. Modifications including 2'-OMe, 5'-NH2, 5'-phosphate, 5'-cholesterol, FAM, Cy3, Cy5, Cy5.5, and biotin can improve stability, silencing efficiency, and visual tracking.
Modified products can be tested for MS and purity as specified in the order, with sequence identity verified.
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  • 6-8business days

    Modified siRNA delivery

  • ≥ 90%

    HPLC purity

  • MSand purity

    Tested as specified in the order

  • 9types

    Common modifications and labels

首页 > Products > siRNA Synthesis > Custom siRNA Synthesis and Modification >
Introduction
Selection Guide
Service Advantages
Service Details
Service Workflow
Delivery Standards
Quality Control Standards
siRNA Performance
Applications
Frequently Asked Questions
Custom siRNA Synthesis and Modification
Chemically Modified siRNA
Fluorescent/Labeled siRNA

Custom siRNA Synthesis and Modification

Comprehensive Options and Flexible Customization

Unmodified siRNA is susceptible to nuclease degradation, has a relatively short serum half-life, and lacks functional labels. Chemical modifications and fluorescent labeling can optimize siRNA performance and add detection capabilities.

We provide custom synthesis services for chemically modified and fluorescently labeled siRNA. Modified products can be tested for MS and purity as specified in the order, with sequence accuracy confirmed.

  • Chemical modification: improves stability, uptake, and silencing efficiency
  • Fluorescent/label modification: monitors transfection, tracks localization, and enables affinity purification
  • Other modification types or custom combinations of multiple modifications

Chemically Modified siRNA

Improves stability, uptake, and silencing efficiency

Introduce specific groups at the sugar ring, phosphate backbone, or terminal positions to improve biological stability, pharmacokinetic properties, and silencing activity.

  • 2'-O-methyl: improves nuclease resistance and reduces immunogenicity and seed-region off-target effects
  • 5'-amino: provides a reactive site for subsequent conjugation
  • 5'-phosphate: optimizes antisense-strand loading into RISC and improves silencing efficiency
  • 5'-cholesterol: improves membrane permeability and uptake by difficult-to-transfect cells

Fluorescent/Labeled siRNA

Transfection monitoring, localization tracking, and affinity purification

Fluorescent dyes and affinity tags are usually attached to the 5' or 3' end of the sense strand to minimize interference with antisense-strand loading into RISC.

  • FAM: Ex 495/Em 520, green fluorescence for monitoring transfection efficiency
  • Cy3: Ex 550/Em 570, orange-red fluorescence for colocalization studies
  • Cy5: Ex 649/Em 670, multicolor imaging and FRET
  • Cy5.5: Ex 675/Em 694, near-infrared deep-tissue in vivo imaging
  • Biotin: affinity purification, pull-down, or ELISA detection

Modification Selection Guide

Choose the appropriate modification type and position for the experimental setting
Experimental Setting Recommended Modification Notes
Routine Cell Transfection FAM or Cy3 fluorescent labeling Monitors transfection efficiency without affecting silencing activity
Improved Stability 2'-OMe sugar modification Improves nuclease resistance and extends half-life
Minimized Off-Target Effects 2'-OMe in the antisense-strand seed region Reduces miRNA-like off-target effects while preserving on-target activity
Improved Silencing Efficiency 5'-phosphate Antisense-strand 5'-P promotes RISC loading
Improves cellular uptake 5'-cholesterol conjugation Improves membrane permeability and transfection efficiency
Multicolor/FRET Studies Dual Cy3+Cy5 labeling Suitable for FRET or colocalization studies
Deep-Tissue In Vivo Imaging Cy5.5 near-infrared labeling Deep tissue penetration for in vivo imaging
Affinity Purification/Pull-Down Biotin labeling Streptavidin magnetic-bead pull-down of interacting proteins
Subsequent Conjugation 5'-amino Provides an amino group for subsequent chemical conjugation

Service Advantages

Multiple modification types, flexible combinations, and professional design support
  • Advanced Synthesis Platform · Optional MS Verification
    Modified products can be tested for MS and purity as specified in the order, with sequence accuracy confirmed. HPLC purity is ≥90% with stable batch-to-batch performance.
  • Multiple Modification Types · Flexible Combinations
    Supports four chemical modifications and five fluorescent/label modifications, available individually or in custom combinations.
  • Modification Design Consultation · Technical Support
    Modification type and position are recommended based on experimental goals, with customizable QC items for performance evaluation.
  • Flexible Scale · Fast Delivery
    Available from 5 nmol to milligram scale. Modified siRNA is delivered in 6–8 business days, with tiered discounts for batch synthesis.

Service Details

Chemical modifications, fluorescent labeling, and custom combinations
Category Modification Type Scale Purification Turnaround Notes
Chemical Modification
 2'-OMe From 5 nmol OPC/HPLC 6 business days Improves stability;
reduces off-target effects
5'-NH2 From 5 nmol OPC/HPLC 6 business days Reactive group for conjugation
5'-Phosphate-on From 5 nmol OPC/HPLC 6 business days Promotes RISC loading
5'-Cholesterol From 5 nmol OPC/HPLC 6 business days Improves cellular uptake
Fluorescent Labeling
 FAM From 5 nmol OPC/HPLC 6 business days Green fluorescence;
transfection monitoring
Cy3 From 5 nmol OPC/HPLC 6 business days Orange-red fluorescence;
colocalization
Cy5 From 5 nmol OPC/HPLC 6 business days Far-red fluorescence;
multicolor/FRET
Cy5.5 From 5 nmol OPC/HPLC 6 business days Near-infrared;
in vivo imaging
Biotin From 5 nmol OPC/HPLC 6 business days pull-down/
Affinity purification
Other Customizations
 Other modifications or multi-modification combinations From 5 nmol OPC/HPLC 6–8 business days Customized to customer requirements

Service note:Modified products can be tested for MS and purity as specified in the order, with sequence accuracy confirmed. Silencing performance should be evaluated in the context of the experimental conditions. For candidate-sequence screening, the three-candidate siRNA package is recommended together with modification add-on services.

Service Workflow

Four steps from modification-design confirmation to synthesis, QC, and delivery
  • Submit Requirements
    Provide the siRNA sequence or target-gene information, modification type and position, scale, and purification method.
  • Confirm the Plan
    Receive the modification plan, molecular-weight calculation, timeline, and quotation within one business day.
  • Synthesis and Quality Control
    Complete modified-strand synthesis, annealing, purification, and optional MS verification, and issue a QC report.
  • Delivery and Support
    Ship lyophilized product with reports and provide performance consultation and transfection-condition recommendations.

Delivery Standards

Complete delivery of modification information and QC reports
  • Lyophilized siRNA
    Aliquoted at 1 OD; store at -20°C protected from light and reconstitute before use.
  • RNA Synthesis Report
    Includes sequence, modification type and position, molecular weight, purification method, and OD value/concentration.
  • Mass Spectrometry Report (Optional)
    Optional MS-spectrum verification confirms the molecular weight of the synthesized and modified sequence.
  • Purity Test Report
    HPLC-purified products include a chromatogram and have a purity of ≥90%.
  • Instructions for Use
    Reconstitution method, storage recommendations, and reference transfection conditions.

Quality Control Standards

Comprehensive QC for modification sites, purity, duplex formation, and appearance
QC Item Test Method Acceptance Criteria
Modification-Site Accuracy Molecular-weight determination by mass spectrometry (MS) (optional) Deviation between measured and theoretical modified molecular weight ≤0.1%
Purity HPLC/OPC OPC:≥75%; HPLC:≥90%
Duplex Formation Native PAGE/CGE Duplex proportion ≥85%
Appearance Visual Inspection White or off-white lyophilized powder

How Do Modifications Affect siRNA Performance?

From stability and delivery to detection and purification, different chemical modifications optimize siRNA performance across multiple dimensions or add detectable functionality
01. Stability and Activity Optimization

Chemical modifications improve stability and optimize RISC loading:

  • 2'-OMe modification:Improves nuclease resistance, reduces immunogenicity and miRNA-like off-target effects, while preserving on-target activity as much as possible
  • 5'-phosphate:Promotes Argonaute recognition and RISC loading, bypassing the rate-limiting intracellular phosphorylation step to improve silencing efficiency
02. Improved Delivery and Cellular Uptake

Conjugated groups improve membrane affinity and cellular uptake:

  • 5'-cholesterol:Uses hydrophobicity to improve membrane affinity and endocytic uptake, enabling some cellular entry even without a transfection reagent
03. Detection, Tracking, and Purification Functions

Labels enable visual monitoring and affinity purification:

  • Fluorescent labels (FAM/Cy3/Cy5/Cy5.5):Used for monitoring transfection efficiency, cellular localization, multicolor colocalization/FRET, and in vivo imaging
  • Biotin：Used in pull-down assays to capture RISC or RNA-binding proteins and in ELISA-like quantitative detection, together with streptavidin-based colorimetric or fluorescence detection

Applications

Covering silencing optimization, experimental quality control, and mechanistic interaction studies

Optimizing Knockdown and Stability

Use 5'-phosphate, 2'-OMe, and 5'-cholesterol to improve RISC loading, nuclease resistance, and uptake by difficult-to-transfect cells.

Experimental Quality Control and Visualization

Monitor transfection efficiency with FAM/Cy3 and track cellular or in vivo distribution dynamically with Cy5.5.

Mechanistic and Interaction Studies

Conduct FRET studies with dual Cy3+Cy5 labeling or capture siRNA-interacting proteins using biotin pull-down.

Frequently Asked Questions

Frequently asked questions about siRNA modification, labeling, and experimental applications
Does siRNA Have to Be Modified?
Standard cell lines such as HEK293 and HeLa can generally use unmodified siRNA directly.
Modification is recommended when monitoring transfection efficiency (fluorescent labeling), improving stability (2'-OMe), enhancing silencing efficiency (5'-phosphate), or promoting uptake (cholesterol). At minimum, a fluorescently labeled control is recommended for transfection monitoring.
What Are the Functions and Recommended Positions of 2'-OMe Modification?
Function: improves nuclease resistance and reduces seed-region off-target effects.
Position: the sense-strand terminus to improve stability, or positions 2–8 in the antisense-strand seed region to reduce off-target effects. Avoid positions 9–11 near the antisense-strand cleavage site.
Is 5'-Phosphate Modification Required?
Not necessarily. Endogenous kinases in standard cells can complete phosphorylation.
It is recommended for rapid onset, low-concentration experiments, or cells with low kinase activity such as primary or stem cells, to support efficient RISC loading.
Does Fluorescent Labeling Affect Silencing Efficiency?
Labeling at the 3' or 5' end of the sense strand has little effect because the sense strand does not mediate cleavage and is discarded after RISC loading. Avoid labeling the antisense strand, which can interfere with loading and cleavage activity.
How Should I Choose Among FAM, Cy3, Cy5, and Cy5.5?
· Choose FAM for routine transfection monitoring (green and economical);
· Choose a Cy3+Cy5 pair for multicolor colocalization or FRET studies;
· Choose Cy5.5 for deep-tissue in vivo imaging (near-infrared).
Confirm that the instrument has the appropriate detection channel before selecting a label.
What Is the Purpose of 5'-Cholesterol Modification?
· Improves cell-membrane affinity and promotes endocytic uptake;
· Suitable for cells sensitive to transfection reagents or uptake-mechanism studies without a transfection reagent;
· Conjugation slightly reduces solubility, so mix thoroughly during reconstitution.
How Is Biotin Labeling Used?
The extremely high affinity of biotin-streptavidin (Kd≈10⁻¹⁵ M) is used to immobilize biotin-siRNA on magnetic beads and pull down interacting proteins, such as RISC components, from lysates for identification by Western blot or mass spectrometry. Biotin is nonfluorescent and requires HRP- or fluorescently labeled streptavidin for detection.
Do Modifications Reduce Silencing Efficiency?
Properly designed modifications have little impact; sense-strand fluorescent labeling and 2'-OMe can retain >90% activity. Incorrect placement, such as modification at the cleavage site or antisense-strand labeling, can significantly reduce efficiency. Validate activity with the unmodified sequence first, then add modifications. Modification-design consultation and customized QC are available.

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