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GoTo DsiRNA Validated Control DsiRNA Sequences
GoTo Bio-Rad siLentMer Validated Target Specific DsiRNA duplexes
View Dicer-substrate Publications


DsiRNA Dicer-substrate siRNAs (DsiRNAs)

Predesigned DsiRNAs - DsiRNA RefSeq Library
 
Over 3,000,000 DsiRNAs have been designed against the approximately 25,000 genes from each of the human, mouse, rat, cow, dog, chicken, and chimp transcriptomes in The fRefSeq Genbank collection: http://www.ncbi.nlm.nih.gov/RefSeq.

Site selection is first performed using a proprietary algorithm that uses novel Dicer-substrate specific design rules. Sequences that pass this stage are next screened to minimize the potential for cross-hybridization and off-target effects (Smith-Waterman analysis); sites are also eliminated that include known SNPs; the analysis is further extended to include the presence of alternatively spliced exons (if any are present for that gene in RefSeq). Finally, local mRNA secondary structure is modeled and areas of high predicted structure are avoided.
 
DsiRNA Libraries
 
  • Splice common: targets all known variants of a gene in RefSeq; duplexes lie only within common exons. This constitutes the bulk of the DsiRNA collection and is the correct choice for most users. 8-10 duplexes are available for each gene target.
  • Splice specific: targets only exons present in specific splice forms. In some cases, the unique exon may be very small or comprise sequence unfavorable for selection of RNAi duplexes (please note that knock-down guarantees do not apply for these sites). This collection is intended for those researchers who have a specialized need to selectively target specific splice variants or isoforms.
When working with a gene target that is included in the above collection, IDT recommends using the pre-designed duplexes as these include significantly more bioinformatics analysis than is possible for sequences designed in real time using the Web interface design tool.
 
Screening DsiRNA
 
Ideal for small scale in vitro applications, screening DsiRNA duplex pricing includes affinity purification. In addition, each duplex is identified by ESI mass spectrometry. Mass spectrometry data is provided free on IDT's website.
 
Screening DsiRNA sequences must be between 24 and 30 bases and 100% complementary with up to a 3-base overhang.

Duplex GuaranteeTube pricePlate Price
2 nmole Screening DsiRNA Duplex$95.00 USD / Duplex$75.00 USD / Duplex
10 nmole Screening DsiRNA Duplex$145.00 USD / Duplex$115.00 USD / Duplex
40 nmole Screening DsiRNA Duplex$225.00 USD / DuplexN/A


Note: The DsiRNA pricing above is for unmodified duplexes only. RNA duplexes can be ordered through the IDT RNA ordering module as ssRNAs or as dsRNA with a variety of modifications.
 


siLentMer Validated Target-Specific DsiRNA
IDT and Bio-Rad have partnered to develop a collection of validated, premade Dicer-substrate duplexes. Hundreds of human genes are currently in the process of validation. Please check the Bio-Rad website for the precise targets available today; note that new products will be introduced rapidly as validation proceeds.
 
Validation criteria require that each duplex reduces target gene expression by >85% at a dose of 5 nM as assessed by qRT-PCR 24 hours post transfection. No other validated reagents must pass such strict criteria. Validated primers for SYBR Green qRT-PCR assays are also available for all targets.
 
See www.bio-rad.com for details and pricing.
 
The Dicector™ Premade DsiRNA Libraries
 
Premade collections of DsiRNAs are available for use in large scale screening programs. Duplexes are arrayed in plates with either 2 nmoles 0.25 nmoles of each duplex. Please inquire for other yield options and custom plate configurations.
Four libraries are currently available. Please inquire for pricing or custom synthesis of new collections of any size.
 
Human Protein Kinome Set
549 Human Protein Kinases, 4 duplexes per target, 2196 duplexes
 
Human GPCR Set
409 Human GPCRs, 4 duplexes per target, 1636 duplexes.
 
Human ATPase Set
78 Human ATPases, 4 duplexes per target, 312 duplexes.
 
Mouse GPCR Set
386 Mouse GPCRs, 4 duplexes per target, 1584 duplexes.
 
Large-Scale DsiRNAs
 
RNA duplexes are available up to 10 grams. All duplexes are purified using RNase-free HPLC methods and come with purity and ESI-MS QC documents.
 
Researchers planning in vivo work should be aware that potentially toxic substances can be introduced during oligonucleotide synthesis and purification (HPLC) processes that must be removed to prevent errors or artifacts.
 
Proper manufacturing protocols for large scale siRNA synthesis include:
  • Adding trace EDTA in HPLC buffers to remove heavy metal cations.
  • Using only biocompatible buffers and salts such as sodium phosphate and sodium chloride for HPLC
  • Maintaining an endotoxin free environment.
  • Options for sterile filtration and endotoxin testing.
 
The Endotoxin Problem
 
The classic endotoxin is lipopolysaccharide (LPS), a natural product present in the outer membrane of the cell wall of Gram negative bacteria. Toxic LPS is liberated whenever these bacteria die. Its presence can cause severe inflammatory responses and/or death of the animals. In mammals LPS binds to serum proteins and interacts with various receptors on monocytes, macrophages, and endothelial cells. This eventually triggers cytokine production, complement activation, and coagulation cascades.
 
IDT large-scale siRNAs are manufactured with in vivo use in mind. Protocols are employed that minimize the risk of endotoxin contamination.
 
As an option, duplexes can be ordered as sterile filtered, endotoxin-free. Endotoxin testing will be performed to verify that levels do not exceed 0.1 EU/mg.
 
Please inquire with IDT Customer Support to obtain a custom quote at 800-328-2661.
 
Biotechniques Protocol Guide to producing siRNA for in vivo applications
 
Dicer-substrate RNAi methods were developed in a collaborative project between IDT and Drs. John Rossi and Dongho Kim at the Beckman Research Institute of the City of Hope National Medical Center.1,2 IDT is licensed under patent applications jointly filed by IDT and The City of Hope Medical Center to sell research products incorporating Dicer-substrate RNAi technology.
 


References

1. Synthetic dsRNA Dicer-substrates enhance RNAi potency and efficacy. Kim, D.H., et al., Nat Biotechnol, 23(2): p. 222-6 (2005).

2. Functional polarity is introduced by Dicer processing of short substrate RNAs. Rose, S.D., et al., Nucleic Acids Res, 33(13): p. 4140-56 (2005).


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