H1 Promoter: A distinct Advantage for shRNA and microRNA
The pSUPER suite of vectors offer the freedom of H1 and the power of Pol III for shRNA expression.
H1 vs. U6
A key component of the pSUPER RNAi System is the H1 RNA polymerase III promoter, which drives the endogenous production of shRNA. While other RNAi vector systems employ U6 promoters, research by the Netherlands Cancer Institute shows H1 to have distinct advantages over other pol III promoters.
Advantages of the Pol III Promoter
Unlike the majority of RNA pol III promoters, type III RNA pol III promoters do not require additional sequence elements and therefore are good candidates for in vivo delivery systems of siRNA. The most abundant cellular RNAs transcribed from a type III RNA pol III promoter are the U6 small nuclear RNA (which play a crucial role in the processing of premature RNA); the 7SK RNA (a negative regulator of RNA polymerase II elongation factor TEFb); and H1 RNA (a component of RNAse P).
The H1 Advantage
The potential negative effects of four different RNA polymerase III promoters on the expression of a heterologous gene driven by a polymerase II promoter were evaluated. The human H1and 7SK promoters displayed no non-specific interferences, whereas the murine and to a lesser extent the human U6 promoters highly interfered with the expression of LGFP. The toxicity of the murine U6 promoter has been previously identified.
Additionally, the H1 promoter proved to offer more flexibility for shRNA target design. In most cases, the +1 nucleotide of the U6-like promoters is always guanosine, whereas the +1 for H1 promoters is adenosine. Interestingly, changing the +1 adenosine to uridine, cytidine, or guanosine within H1-expressed stem-loop sequences does not seen to affect gene silencing, indicating that H1 promoters may be more flexible than U6 promoters in regard to +1 sequence changes1.
Therefore, the human H1 promoter was used in subsequent experiments aimed to evaluate the gene silencing potential of the pSUPER RNAi System, with highly successful results2,3.
Stable Suppression by shRNA or microRNA
The pSUPER RNAi System allows researchers to perform long-term experiments to study the effect of blocking the production of specific proteins or microRNA within cells and whole organisms- leading to new therapeutic weapon against many diseases including HIV, cancer and other genetic disorders.
With silencing effects lasting up to two months or more, this new system also unlocks the power of functional genomics for target validation and drug discovery, two of the key components in the gene-to-drug pathway.
The pSUPER Heritage
The original pSUPER vector was first described in a paper published in the prestigious journal Science in April 2002. Such was its impact that an unprecedented 1,500 requests were received from research scientists asking for access to the vector for their own studies - before the vector was even available for sale.
As a result the pSUPER vector is becoming a standard workhorse in laboratories worldwide for shRNA, microRNA, and X-miR MicroRNA repressors, providing insight into the essential genes involved in a wide range of diseases - from neurodegenerative diseases such as Alzheimer's and Parkinson's, through infectious diseases such as HIV and hepatitis, to cancer and other genetically based diseases.
1. Tuschl, T., Nature Biotechnology 20, 446 (2002)
2. T.R. Brummelkamp, R. Bernards, and R Agami, Science 296, 550 (2002).
3. T.R. Brummelkamp, R. Bernards, and R Agami, Cancer Cell 2:3, 243-247 (2002)
Getting Started with pSUPER or pSUPERIOR
The pSUPER RNAi System generates long-term loss-of-function phenotypes in mammalian cell lines by utilizing a novel vector, along with a pair of oligos unique to the target gene that are ligated into the vector, to direct the persistent synthesis of siRNA intracellularly. Follow these guidelines when designing your oligos:
A guide to design 60-mer shRNA for pSUPER. shRNA is designed automatically in Oligoengine 2.0.
Targeting Your Gene with Asymmetric shRNA or X-miR
The pSUPER vector requires the insertion of two annealed long-mer oligos. We have pre-designed thousands of Asymmetric shRNA, microRNA clones, or X-miR MicroRNA Repressors. Please email us for shRNA design specific to your project or check online in our store.
shRNA, microRNA repressors, or microRNA for pSUPER contain the ends compatible with digestion using either BglII/HindIII (default) or BglII/XhoI.
pSUPER clones expressing all known mature microRNA are available in our store
pSUPER.retro and Packaging Cells
Oligoengine recommends the use of the Phi-NX cell line, a 293T-based line developed by the Nolan Lab at Stanford. The Phi-NX cell line is commercially available at AGCC. Alternatively, other cell lines have proven successful with pSUPER.retro. Phe-NX Cell Line
pSUPERIOR and Inducible RNAi
With the use of a TETR binding protein, pSUPERIOR gives you the option of controlling suppression inducibly with Tetracycline or Doxycycline.. pcDNA6/TR Information
All pSUPER and pSUPERIOR vectors are available exclusively in circular form.
Any Target + pSUPER or pSUPERIOR + 18 Days.
Pick any pSUPER or pSUPERIOR vector, a target gene, and we will send a sequenced clones for $599 each. When you order three, we will guarantee suppression greater than 70%.
Cloning Service Deliverables:
• Clones can express either Asymmetric shRNA, mature microRNA, or X-miR MicroRNA Repressors.
• All clones are sequence confirmed.
• 18 Business Day turnaround.
• Price includes all materials.
• Delivered as miniprep, ~4 µg.
Published Papers Using pSUPER.
We are amassing a list of hundreds of published papers which successfully utilize pSUPER vectors.
Get your paper included.
Email customer service with your reference for using pSUPER and we will be happy to cite your work on the site.
pSUPER RNAi System Protocols
Click to download a PDF of the pSUPER or pSUPERIOR protocol.
pSUPER RNAi System Maps
Click to download a PDF of the pSUPER or pSUPERIOR maps.
pSUPER RNAi System Sequences
Click to download a text file of the pSUPER or pSUPERIOR sequences.
pSUPER.retro.puro (Stuffered Version) Sequence:
Licensing the pSUPER RNAi System.
Now, to better support the needs of our commercial customers, individual pSUPER vectors are available to non-academic/for profit organizations under more flexible and affordable licensing terms.
Academic and Non-Profit Licensing Terms
Researchers at academic and non-profit institutions are granted a limited end-user license with the purchase of each pSUPER vector.
Commercial and For-Profit Licensing Terms
Previously, use of the pSUPER RNAi System by commercial pharma and biotech companies has been subject to a comprehensive licensing program. Now companies can purchase individual pSUPER vectors under a more streamlined product use agreement. Additionally, any products, including cell lines, transgenic animals and therapeutic compounds developed using the pSUPER RNAi System will be considered as licensed products and subject to a sales royalty by Cancer Research Technologies.
For more information, please contact an OligoEngine Customer Service.