Custom siRNA Synthesis

Custom siRNA Synthesis

siRNA customization has never been easier. Numerous combinations of modifications, sizes, and purification options are available for convenient online ordering.

Custom siRNA Synthesis Overview

Custom siRNA synthesis options
Custom siRNA Order
Modified siRNA synthesis Order
Multiple siRNA synthesis Order
Custom SMARTpool design & synthesis Order
siRNA Design Tools
siDESIGN Center Design Now

Learn more about our proprietary siRNA modifications, dye labels, other chemical modifications, purification and processing options and in vivo RNAi by browsing the tabs at the top of this page.

Choose from a variety of custom options to meet your experimental needs

siRNA customization has never been easier. Numerous combinations of modifications, synthesis scales and purification options are available for convenient online ordering.

Consult the siRNA yield table for estimated yield information on unmodified siRNA at various synthesis scales.

Expert siRNA design assistance is available

If you are working in human, mouse or rat models, we may already have what you need as a pre-designed product: Search for your gene in the Search field found in the upper right corner.

Use the siDESIGN Center, a free online siRNA design tool, to design siRNAs targeting genes in non-standard species, particular splice variants or homologous regions across gene families or species.

Take advantage of our internal design experts and SMARTpool technology! Order a functionally guaranteed Custom SMARTpool for worry-free customization.

Our capabilities include a range of alternative siRNA designs to empower your RNAi research

If you can't find what you need or you require additional specifications not available through online order, please request a quote. Alternative siRNA designs that we support include:

  • Blunt-ended
  • Asymmetrical
  • Longer duplexes (>23 nt)
  • Mismatched strands
  • Alternative bases or linkages

If you would like further assistance, please contact our experienced team of Technical Support Scientists at +1 800 235 9880 or ts.dharmacon@ge.com.

Proprietary siRNA

Modified for specificity, stability or self-delivery, we have the siRNA solution.

Choose one of our specialized chemical modification patterns to enhance your custom siRNA.

These proprietary siRNA modifications are available only with Dharmacon pre-designed products or custom siRNA Synthesis. Additional modifications (dyes, linkers, etc.) are available via the Modified siRNA Synthesis ordering page.

  • ON-TARGET: Enhanced antisense (guide) strand loading into RISC
  • ON-TARGETplus: Reduces off-target activity from both strands for premium specificity
  • Accell: Delivers siRNA into difficult-to-transfect cells without a transfection reagent
  • siSTABLE: Greater stability in nuclease-rich environments for in vivo applications
ON-TARGET ON-TARGETplus Accell siSTABLE
Inhibits sense (passenger) strand uptake by RISC ON-TARGET siRNA Inhibits RISC ON-TARGET Plus siRNA Inhibits RISC Accell siRNA Inhibits RISC siSTABLE siRNA Inhibits RISC
Antisense strand seed region modified for greater specificity to target ON-TARGET Plus siRNA Antisense strand
Resistant to endo-and exonuclease degradation Accell siRNA Resistant to endo-and exonuclease degradation siSTABLE siRNA Resistant to endo-and exonuclease degradation
Delivery into cells without transfection reagent Accell siRNA without transfection reagent
Also available as Pre-designed siRNA ON-TARGET Plus siRNA Pre-designed Accell siRNA Pre-designed
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On-Target siRNA antisense strand bias

ON-TARGET

for ensuring antisense strand bias

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ON-TARGET Plus siRNA premium specificity

ON-TARGETplus

for premium specificity

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Accell siRNA difficult-to-transfect cells

Accell

for difficult-to-transfect cells

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siSTABLE siRNA nuclease resistance

siSTABLE

for nuclease resistance

Dye Labels

Dharmacon offers a broad portfolio of dye modifications for fluorescent labeling of your custom siRNA and RNA oligonucleotides.

  • Include a fluorescent tag at the 3' or 5' positions on either strand of an siRNA
  • Photostability of DyLight dyes from pH 4-9 frequently exceeds that of Cy dyes, with comparable spectra
  • Synthesis of most dye-labeled siRNAs is scalable to gram quantities

Fluorescent dyes available online:

*Some 3' dye labels are only available upon request.
Fluorophore λ max abs (nm) λ max em (nm) Comparable to 5' or 3'
Fluorescein / 6-FAM 494 520 - both*
DY547 548 562 Cy3, Alexa 546, Alexa 555 both*
TAMRA 565 580 - both*
DY647 645 662 Cy5, Alexa 647 both*
DY677 684 698 CY5.5, Alexa 680 both*

Some of the dye options listed in the above tables will produce higher oligo yields than others. For assistance in choosing the most appropriate dye label for your application, please contact a Technical Support Scientist at +1 800 235 9880 or ts.dharmacon@ge.com. You can also request a quote online if you already know that an alternative dye will be required for your experiments.

Alexa Fluor® is a registered trademark of Invitrogen Corporation. All other trademarks are the property of GE Healthcare Limited and its subsidiaries.

Chemical modifications available for custom siRNA

In addition to fluorescent dyes, we offer a broad portfolio of other chemical modifications that can also be applied to both siRNA and single-stranded RNA.

For information on our specialized modification patterns for reducing off-targets, self-delivery and nuclease resistance, view the Proprietary siRNA tab.

Modified siRNA

siRNA can be modified at either end, on either strand. Multiple modifications are also available but availability may be dependent upon sequence or other factors. Please contact Technical Support for more information

The following standard chemical modifications are available for online order:

5' Modifications 3' Modifications
TAMRA Biotin
Phosphate idT (inverted deoxythymidine)
N6 (amino, 6-carbon) linker Puromycin
Biotin ddC (terminal dideoxy-cytidine)
Fluorescein Fluorescein
Cholesterol Cholesterol
Disulfide Thiol-Modifier N6 (amino, 6-carbon) linker
Dy547 Dy547
Dy647
Dy677
Tetrachloro-fluorescein (TET)

Standard modifications for single-stranded RNA can also be applied to siRNA by request. Please contact Technical Support to inquire about additional modifications or options for your siRNA, or request a quote now.

Purification & Processing

We routinely achieve 80-85% purity for unmodified siRNA strands without additional purification.

However, purification may be recommended when chemically synthesized siRNAs are:

  • Chemically modified at the 3' end or internally
  • Dually modified on the same strand
  • Intended for use in highly sensitive assays or in vivo applications

Please contact Technical Support for information regarding recommendations for purification or purity estimates for unpurified material at +1 800 235 9880 or ts.dharmacon@ge.com.

siRNA Processing Options:

Single Strands (A1) Standard (A4) PAGE (C) HPLC In vivo In vivo HPLC
Desalted Desalted Standard siRNA desalted PAGE siRNA HPLC siRNA Desalted in vivo siRNA Desalted In vivo HPLC siRNA Desalted
Deprotected Deprotected Standard siRNA Deprotected PAGE siRNA Deprotected HPLC siRNA Deprotected in vivo siRNA Deprotected in vivo HPLC siRNA
Duplexed Duplexed Standard siRNA Duplexed PAGE siRNA Duplexed HPLC siRNA Duplexed in vivo siRNA Duplexed in vivo HPLC siRNA
Purified Purified PAGE siRNA Purified HPLC siRNA in vivo HPLC siRNA
Endotoxin tested Endotoxin tested in vivo siRNA Endotoxin tested in vivo HPLC siRNA
Sodium counter-ion exchange Sodium counter-ion exchange in vivo siRNA Sodium counter-ion exchange in vivo HPLC siRNA
Recommended for modified siRNA (dyes, etc.) Recommended PAGE Modified siRNA Recommended HPLC Modified siRNA Recommended in vivo HPLC siRNA
Recommended for in vivo use Recommended for in vivo sirna Recommended for in vivo HPLC siRNA
2'ACE protected single- strands 2 ACE protected single strands siRNA

Description of siRNA Processing Terms

  • Desalted: The siRNA duplex has been desalted by either ethanol precipitation or C18 column desalting
  • Deprotected: The 2'-ACE protecting groups of the RNA bases have been removed (deprotected)
  • Duplexed: The two complementary siRNA strands have been annealed to form a duplex
  • Single-stranded (A1): The siRNA is provided with the sense and antisense strands in separate tubes; individual strands have NOT been desalted or deprotected
  • Standard (A4): The siRNA is provided as a desalted & deprotected duplex, ready to use upon resuspension
  • PAGE: The siRNA duplex has been purified by Polyacrylamide Gel Electrophoresis
  • HPLC: The duplex has undergone ion exchange High Performance Liquid Chromatography for purification
  • In vivo: The duplex has been processed by counter-ion (Na+) exchange, desalting, sterile filtration, and endotoxin testing
  • In vivo HPLC: The duplex has undergone both in vivo processing as well as HPLC purification

In vivo RNAi

Partner with the industry leader in RNAi technologies for the highest quality in vivo -ready RNAi reagents.

We want to ensure your in vivo experiments have the best chance of success. To assist you in your experimental planning, we offer the following options and guidance:

Assess your siRNA sequences in vitro

  • We recommend identifying a highly functional siRNA design in vitro prior to initiating expensive experiments using animal models
  • A predesigned Set of 4 siRNA reagents is ideal for testing multiple siRNA sequences

Consider a proprietary siRNA modification pattern for enhanced nuclease resistance

  • siSTABLE modifications prevent degradation from exo-and endonucleases. siSTABLE is recommended when the siRNA will be exposed to a biological environment rich in nucleases, such as animal serum.
  • Accell modifications incorporate delivery-enhancing properties in addition to stability enhancement. Accell is recommended when target cell or tissue types are not amenable to standard modes of delivery.
  • See a list of recent publications using siSTABLE and Accell modified siRNA for in vivo experimentation.

ORDER custom siRNA with siSTABLE or Accell

Carefully calculate the total siRNA quantity required

  • Consider the number of animals in each cohort, the number of treatments or doses that will be required, and the amount of siRNA required for each dose
  • In vivo experiments typically require a large quantity of siRNA. Synthesis capabilities can accommodate up to 10 grams by request, or up to 100 mg available online
  • Need help converting nmol to mg? Check our siRNA yield table.

Consider in vivo processing of your siRNA when toxicity to the animal is of concern

  • A specialized post-synthesis siRNA processing procedure including counter-ion (Na+) exchange, sterile filtration, desalting, and endotoxin testing
  • Available with or without HPLC purification. For more information, review our siRNA purification and processing options.
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Accell siRNA

Accell

for difficult-to-transfect cells

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siSTABLE nuclease resistance

siSTABLE

for nuclease resistance

For more information on important considerations for in vivo experimentation see our Technical Note: "In vivo RNAi: Biodistribution, Delivery, and Applications"

ON-TARGET modification ensures antisense strand bias


ON-TARGET Modification antisense strand bias

The ON-TARGET modification promotes correct strand uptake by blocking the sense (passenger) strand from being taken up by the RISC process. This is a useful method for ensuring antisense (guide) strand processing.

All proprietary siRNA modifications (ON-TARGETplus, Accell, siSTABLE) incorporate this sense-strand modification to promote antisense (guide) strand-facilitated silencing.

ON-TARGETplus siRNA dual-strand modification pattern for reduction of off-targets


ON-TARGET Plus dual-strand modification

A 2006 publication demonstrates that off-target effects are primarily driven by antisense strand seed activity.1 Therefore, sense strand inactivation alone does not decrease the total number of off-target genes. ON-TARGETplus modifications account for both strands:

  • Sense strand is modified to prevent interaction with RISC and favor antisense strand uptake
  • Antisense strand seed region is modified to minimize seed-related off-targeting

The ON-TARGETplus modification pattern dramatically reduces off-targets. Off-target effects induced by the indicated siRNAs were quantified using microarray analysis. For each target, three different siRNAs were used: unmodified, sense strand-inactivated, and ON TARGETplus-modified. Data shown represents genes down-regulated by two-fold or more. HEK293 cells were transfected with 100 nM siRNA using 0.2 µL of DharmaFECT 1. Data was analyzed at 24 hours.

1 Jackson, A.L. et al. "Position-specific Chemical Modification Increases Specificity of siRNA-mediated Gene Silencing." RNA 12.7 (2006) 1197-1205.

Accell delivery and gene silencing in cardiomyocytes


Accell delivery and gene silencing cardiomyocytes

Neonatal rat ventricular myocytes were incubated with 1 µM Accell Green (A; Cat# D-001950-01) or Red (B; Cat# D-001960-01) Non-targeting siRNA for 72 hours in Accell delivery media (Cat# B-005000). Nuclei were stained with DAPI (blue). Labeled control uptake showed diffuse cytoplasmic localization in nearly all cells.

The bar graph indicates the level of gene silencing achieved with Accell GAPD Control siRNA (Cat# D-001930-03) and Pool (Cat# D-001930-30) control reagents when used with neonatal rat ventricular myocyte (NRVM) media or Accell delivery media.

Myocytes were prepared as described in Maass AH & Buvoli M. Cardiomyocyte preparation, culture, and gene transfer. Methods Mol Biol. 2007;366: 321-30. mRNA expression was determined by QuantiGene branched DNA assay (Panomics).

siSTABLE-modified siRNAs resist degradation by nucleases


siSTABLE modified siRNAs resist degradation by nucleases

Conventional siRNA is degraded within minutes in serum-containing environments, making in vivo use of siRNA problematic. This graph presents evidence that the siSTABLE modification pattern dramatically extends the half-life of siRNA in the presence of 100% human serum as compared to Stealth RNAi (Invitrogen).

Accell siRNA also includes these stability-enhancing modifications, as well as providing delivery to difficult-to-transfect cells without a transfection reagent.

5 Fluorescein

5'-Fluorescein

Description: Fluorescein is often used in fluorescence experiments to demonstrate the kinetics of folding or substrate binding. Fluorescein is also used as a donor to track optimal changes related to folding or substrate binding to intermolecular interactions.

  • Mol. Wt.: 537.46
  • Special Note: Light sensitive. Fluorescein is very sensitive to photo bleaching.
  • Absorbance/Fluorescence: 494/520 nm

Reference: Science 266: 785-789 (1994), EMBO J. 17: 2378-2391 (1998)

5 DY547 CY3 Alternate

5'-DY547 (Cy3™ alternate)

For more information please contact Technical Support:

ts.dharmacon@ge.com
1-800-235-9880 (US)
00800-222 00 888
+44 (0) 845 3 63 04 25 (UK / EMEA)

  • Mol. Wt.: 491.25 g/mol
3'-DY547

3'-DY547 (Cy3™ alternate)

For more information please contact Technical Support:

ts.dharmacon@ge.com
1-800-235-9880 (US)
00800-222 00 888
+44 (0) 845 3 63 04 25 (UK / EMEA)

  • Mol. Wt.: 563.30 g/mol
5 Tamra-hexyl Linker

5'-TAMRA-hexyl linker

Description: TAMRA is a strongly absorbing dye with a wide variety of applications. This modification is coupled from the 5'- or 3'-end of an oligonucleotide to either the 5th or 6th position of the dye.

  • Special Note: Light sensitive
  • Absorbance/Fluorescence: 565/580 nm

References: Nucl. Acids. Res. 24: 4535-4542 (1996), Biochem. 39: 14487-14484 (2000)

5 DY647 CY5 Alternate

5'-DY647 (Cy5™ alternate)

For more information please contact Technical Support:

ts.dharmacon@ge.com
1-800-235-9880 (US)
00800-222 00 888
+44 (0) 845 3 63 04 25 (UK / EMEA)

  • Mol Wt.: 517.26 g/mol
5 Dy677 CY5.5 Alternate

5'-DY677 (Cy5.5™ alternate)

For more information please contact Technical Support:

ts.dharmacon@ge.com
1-800-235-9880 (US)
00800-222 00 888
+44 (0) 845 3 63 04 25 (UK / EMEA)

  • Mol Wt.: 617.29 g/mol

What is an NHS Ester?

An NHS ester is a reactive group found on the dye that provides the functionality for labeling amino groups found on the oligonucleotide.

Some DyLight dyes are available as phosphoramidites (our standard dyes), and others as NHS esters (our non-standard Alexa alternatives). If applicable, we recommend DyLight dyes as a first choice since this form is most compatible with our 2'-ACE RNA synthesis chemistry and will therefore be more likely to produce higher yields.

siRNA Yield Table


For unmodified siRNA, the following approximate yields can be expected:
Standard (A4) Purified in vivo in vivo HPLC
nmol mg nmol mg nmol mg nmol mg
0.025 µmol scale 20 0.25 - - - - - -
0.05 µmol scale 40 0.5 20 0.25 25 .3 - -
0.2 µmol scale 150 2 80 1 100 1.3 50 0.65
0.4 µmol scale 300 4 160 2 200 2.6 100 1.3
1.0 µmol scale 750 10 320 4 500 6.6 250 3.3
2.0 µmol scale 1500 20 750 10 1000 13 500 6.6
5.0 µmol scale 3750 50 1875 25 2500 33 1250 16
10.0 µmol scale 7500 100 3750 50 5000 66 2500 33
Pre-designed siRNAs are available in the following formats:
  • Individual duplexes: Four individual siRNAs are pre-designed for every gene in human, mouse and rat. Order 1, 2, 3 or 4.
  • SMARTpool: All four siRNAs are pooled together as a single reagent in one tube.
  • Set of Four: All four siRNAs are provided as individual duplexes in four individual tubes.

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