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Internal Oligo Modifications for DNA & RNA Probes

Custom internal amino, thiol, azide, alkyne, fluorescent, quencher, spacer, photocrosslinker, and stability modifications for advanced DNA and RNA oligonucleotide workflows.

Explore related spacer & linker modifications, quencher compatibility, and oligo bioconjugation services.

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Overview Selection Guide Reactive Handles Dyes & Quenchers Spacers Photocrosslinkers Stability Analogs Design & QC Contact
Overview

Internal Modifications for In-Sequence Functionality

Internal oligo modifications add chemistry within the DNA or RNA sequence rather than only at the 5′ or 3′ ends. These in-sequence modifications enable bioconjugation, built-in reporter systems, geometry control, photocrosslinking, and stability tuning for custom probes, ASO, siRNA, aptamer, FRET, qPCR, and structural biology workflows.

Bio-Synthesis offers a broad range of internal oligonucleotide modifications including amino, thiol, azide/alkyne, aldehyde, DBCO, fluorescent bases, dark quenchers, HEG/TEG and alkyl spacers, psoralen, halogenated bases, LNA, 2′-OMe, 2′-F, UNA, and related analogs. If the internal modification you need is not listed below, our scientific team can evaluate custom synthesis, phosphoramidite sourcing, or post-synthetic modification strategies.

Our team can support probe architecture, dye–quencher spacing, click chemistry planning, ASO/siRNA modification placement, purification selection, and ISO-aligned analytical QC including HPLC, ESI-MS, and optional CE/SEC or functional testing.

Click-Ready Handles

FRET / qPCR Reporters

Spacers & Geometry

Photocrosslinking

Tm & Stability

Design & QC Support

Internal Modification Selection Guide

Browse Internal Modifications by Function

Start with the role your internal modification needs to perform, then jump directly to the relevant chemistry, design, or QC category.

⚛

Install a Reactive Handle

Use amino, thiol, azide, alkyne, aldehyde, DBCO, halo, lipid, or serinol handles for ligation and conjugation.

View Reactive Handles →

✦

Add Reporters or Quenchers

Use internal dyes and dark quenchers for qPCR, molecular beacons, FRET probes, multiplex assays, and imaging.

View Dyes & Quenchers →

⟷

Control Spacing & Geometry

Use dSpacer, HEG/TEG, alkyl spacers, and internal linkers to manage reach, sterics, and probe architecture.

View Spacers →

☀

Capture Interactions

Use psoralen, halogenated bases, and thio bases to trap nucleic acid–nucleic acid or nucleic acid–protein contacts.

View Photocrosslinkers →

⇧

Tune Stability & Affinity

Use LNA, BNA, cEt, 2′-OMe, 2′-F, UNA, and TNA analogs to adjust Tm, nuclease resistance, and potency.

View Stability Analogs →

✓

Plan Design & QC

Get support for modification placement, purification strategy, analytical confirmation, documentation, and custom synthesis feasibility.

View Design & QC →

Reactive Handles

Internal Reactive Handles for Conjugation

In-sequence NH2, SH, azide, alkyne, aldehyde, halo, SPAAC, and lipid/serinol handles support post-synthetic ligation of dyes, peptides, haptens, polymers, chelators, and other payloads.

Product Description Typical Use Code
3′-Azido-Modifier Serinol Internal azide serinol handle. CuAAC/SPAAC click with DBCO or alkynes. [INT-N3-Serinol]
Biotin-TEG Azide Biotin tag with TEG-azide spacer. SPAAC to DBCO; capture and immobilization. [Bio-TEG-N3]
Desthiobiotin-TEG Azide Reversible biotin analog with azide. Affinity capture with mild elution. [dBio-TEG-N3]
Psoralen Azide Psoralen headgroup with azide. Click installation followed by UV crosslink mapping. [Psor-N3]
Coumarin Azide Blue/UV fluorophore with azide. Clickable fluorescent labeling. [Coum-N3]
6-FAM-TEG Azide FAM dye with TEG-azide spacer. CuAAC/SPAAC dye attachment. [FAM-TEG-N3]
Disulfo-Cyanine 7 Azide Water-soluble NIR azide dye. Low-background far-red labeling. [sCy7-N3]
TEMPO / TEMPO-TEG Azide Nitroxide spin-label with azide. EPR and paramagnetic probes. [TEMPO-N3]
5′-DBCO-TEG 5′ cyclooctyne with TEG spacer. Copper-free SPAAC to azides. [5′-DBCO-TEG]
DBCO-dT Internal DBCO on thymidine. SPAAC to azides, dyes, or ligands. [INT-DBCO-dT]
C8-Alkyne-dC / C8-Alkyne-dT Internal terminal alkyne on dC or dT. CuAAC click ligation. [C8-C≡C-dN]
5-Ethynyl-dU 5-ethynyl uridine analog. Internal alkyne for click chemistry. [5-Ethynyl-dU]
5′-Aldehyde-Modifier C2 5′ carbonyl handle. Oxime or hydrazone ligation. [5′-CHO-C2]
Amino-Modifier C6 dR Long-arm internal amine. NHS-ester coupling to dyes or peptides. [INT-NH2-C6]
Palmitate Serinol C16 lipid on serinol. Uptake enhancement and anchoring. [Palm-Ser]
Technical notes for internal reactive handles
  • SPAAC with DBCO and azide is copper-free and useful for sensitive biomolecules.
  • For CuAAC workflows, use TBTA/THPTA ligands and remove copper thoroughly by HPLC or cleanup.
  • Use HEG/TEG spacers near bulky cargos to preserve hybridization and reduce quenching.
Fluorescent & Quencher Bases

Internal Fluorescent Labels, Bases & Quenchers

Internal dyes and dark quenchers support qPCR probes, molecular beacons, FRET readouts, multiplex assays, and imaging workflows.

Product Description Typical Use Code
6-FAM-TEG Azide FAM dye with TEG-azide spacer. Clickable dye installation for qPCR/FRET. [FAM-TEG-N3]
6-HEX Azide HEX dye as azide. Clickable green/yellow labeling. [HEX-N3]
6-TET Azide TET/JOE-like dye as azide. Clickable green-yellow labeling. [TET-N3]
Coumarin Azide Blue/UV fluorophore. Environment-sensitive readouts. [Coum-N3]
Cyanine 3 / 3.5 Orange/red-shifted Cy dyes. qPCR and FRET probes. [Cy3|Cy3.5]
Cyanine 5 / 5.5 Far-red and NIR dyes. Low-background far-red channels. [Cy5|Cy5.5]
JOE-dT Internal JOE on dT. Beacon/probe reporter. [JOE-dT]
Quasar® 570-dT Internal orange-red dye. Probe channels and multiplexing. [Q570-dT]
Quasar® 670-dT Internal far-red dye. Far-red probe channels. [Q670-dT]
Thiazole Orange NHS Ester Environment-sensitive dye. Label NH2-oligos for intercalative readouts. [TO-NHS]
BHQ™-1 Non-emissive quencher, 480–580 nm. FAM/HEX/TET probe quenching. [BHQ-1]
BHQ™-2 Non-emissive quencher, 560–670 nm. Cy3/ROX probe quenching. [BHQ-2]
BBQ-650®-dT Internal dark quencher for red/far-red channels. Cy5/Quasar 670 beacons and probes. [BBQ-650-dT]
Technical notes for internal dyes and quenchers
  • Start with 15–25 nt donor–quencher spacing and refine empirically for each probe.
  • Add HEG/TEG spacing near dyes when bulky reporters or quenchers reduce hybridization performance.
  • Prefer dark quenchers such as BHQ/BBQ in multiplex assays to minimize bleed-through.
Spacers & Linkers

Internal Spacers, Linkers & Geometry Modifiers

Internal abasic sites, alkyl spacers, PEG/HEG/TEG linkers, and photocleavable linkers help control geometry, reach, steric spacing, and probe architecture.

Product Description Typical Use Code
1-Ethynyl dSpacer Abasic spacer with terminal alkyne. Internal click site with minimal base perturbation. [dSpacer-C≡CH]
dSpacer / Abasic Site Internal non-base spacer. Geometry control, gap creation, structural studies. [dSpacer]
HEG / TEG Spacer Flexible ethylene-glycol spacer. Reduce steric hindrance near dyes, surfaces, or cargos. [HEG|TEG]
C3–C12 Alkyl Spacers Variable-length alkyl spacers. Adjust reach and separation between functional groups. [C3-C12]
Photocleavable Linker Light-cleavable internal linker. Controlled release and photo-regulated experiments. [PC-Linker]
Technical notes for spacers and linkers
  • Use longer PEG/HEG or C9–C12 spacers for bulky cargos, surfaces, nanoparticles, or protein conjugates.
  • Shorter spacers preserve compact probe architecture but may increase steric conflict.
  • Spacer selection should be optimized with dye, quencher, ligand, or surface geometry in mind.
Photocrosslinkers

Internal Photocrosslinkers for Interaction Mapping

UV-activated bases and crosslinkers can covalently trap DNA/RNA or nucleic acid–protein contacts for structure, binding, and interaction studies.

Product Description Typical Use Code
Psoralen C2 Short-reach psoralen crosslinker. DNA interstrand crosslinks at 320–365 nm. [Psor-C2]
Psoralen C6 Psoralen with longer spacer. Crosslinking opposite strand/base. [Psor-C6]
Psoralen Azide Clickable psoralen headgroup. Site-specific installation followed by UV crosslink. [Psor-N3]
5-I-dT Halogenated iodo-thymidine. UV crosslinking and heavy-atom derivatization. [5-I-dT]
TIPS-5-Ethynyl-dU Protected alkyne dU. Click handle and photoprobe workflows. [TIPS-Eth-dU]
Thio Bases Photoactive sulfur-containing bases. UV-mediated trapping and structural studies. [Thio-N]
Technical notes for photocrosslinking designs
  • Optimize irradiation wavelength, dose, oxygen, and exposure time to balance crosslink yield versus damage.
  • Psoralen is useful for nucleic acid interstrand crosslinking, while halogenated bases support crosslinking and structural work.
  • Plan purification and MS strategy before synthesis because some photoproducts complicate analytical interpretation.
Stability & Sugar Analogs

Internal Stability, Sugar & Affinity Analogs

Locked/bridged bases, 2′-modified sugars, flexible analogs, and XNA-type modifications help tune nuclease resistance, hybridization affinity, safety, and activity.

Product Description Typical Use Code
2′-OMe-N6-Me-A 2′-O-methyl sugar with N6-methyl adenine. Epitranscript mimic; stability and tuning. [2′OMe-m6A]
LNA / BNA / cEt Locked or bridged sugar analogs. Increase Tm, nuclease resistance, and ASO potency. [LNA|BNA|cEt]
2′-OMe / 2′-F Common RNA stabilizing sugar modifications. siRNA, ASO wings, immune profile tuning. [2′OMe|2′F]
UNA / TNA Flexible or alternative nucleic acid frameworks. Fine-tune duplex behavior and reduce over-stabilization. [UNA|TNA]
Technical notes for stability analogs
  • For gapmer ASOs, keep locked bases in the wings and maintain a DNA-like RNase H core.
  • 2′-OMe and 2′-F patterns are commonly used to improve siRNA stability and reduce immune stimulation.
  • UNA can reduce Tm and may help tune overly stable probes or reduce aggregation.
Design & QC

Design Strategy, Synthesis & Analytical Support

🧪

Design Consulting

  • Probe architecture and dye–quencher spacing.
  • Click chemistry planning for azide, alkyne, DBCO, amine, and thiol handles.
  • ASO, siRNA, gapmer, and reporter placement guidance.
⚙

Synthesis & Scale

  • Scalable synthesis from nmol to gram-level projects.
  • Support for difficult sequences, bulky internal modifications, and custom phosphoramidites.
  • Optional post-synthetic conjugation and cleanup workflows.
✓

QC & Documentation

  • HPLC purity and ESI-MS identity confirmation.
  • Optional CE, SEC, photochemistry, or functional testing.
  • ISO-aligned documentation and SKU mapping available.
Quality alignment: Bio-Synthesis supports ISO 9001 / ISO 13485-aligned workflows and GLP/GMP-like practices as scoped.

Frequently Asked Questions

FAQ

What are internal oligo modifications?
Internal oligo modifications add functional chemistry within the sequence, including amino, thiol, azide, alkyne, aldehyde handles, fluorescent bases, quenchers, spacers, linkers, photocrosslinkers, and stability analogs.
What QC is provided for custom internal oligonucleotides?
Standard QC includes HPLC purity and ESI-MS identity. Optional CE, SEC, photochemistry, functional testing, and ISO-aligned documentation can be added depending on project requirements.
What are photocrosslinking oligonucleotides?
Photocrosslinking oligos use psoralen, 5-iodo-dT, thio bases, or related photoactive chemistries to covalently trap nucleic acid–nucleic acid or nucleic acid–protein contacts after UV activation.
Can internal modifications improve siRNA and ASO stability?
Yes. LNA/BNA/cEt, 2′-OMe, and 2′-F can raise Tm and improve nuclease resistance. For gapmers, preserve the DNA-like core for RNase H and keep stabilizing modifications in the wings.
How are amino, thiol, and azide modifications used in DNA probes?
Amino groups couple to NHS esters, thiols bind maleimides or gold surfaces, and azide/alkyne handles support SPAAC or CuAAC click chemistry. Spacers are often added to reduce steric effects.
Which internal fluorescent bases are useful for qPCR and FRET?
FAM/BHQ-1 and Cy3/BHQ-2 are common starting pairs, while Cy5/BBQ-650 can support far-red designs. Donor–quencher spacing and spacer placement should be optimized for each probe.
More FAQ'sAll FAQ's
Contact & Quote Request

Need help designing an internal-modified oligo?

For the fastest review, share your oligo type, sequence, internal modification position, reactive handle or reporter, scale, purification target, conjugation needs, QC requirements, and intended application. Our scientific team can recommend the most suitable internal modification strategy and provide a detailed quote.
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