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pSCREEN-iT™/lacZ-DEST Gateway™ Vector Kit Invitrogen™

Using the pSCREEN-iT™/lacZ-DEST Gateway® Vector Kit, you can:

•Assess the potency of any RNAi knockdown reagent using a fast, sensitive, and cost-effective β-gal reporter assay
•Identify the best knockdown reagent for your RNAi experiments without knowing protein function, western blotting, or qRT-PCR

How it Works: The BLOCK-iT™ RNAi Target Screening system method uses a β-galactosidase activity assay to measure the knockdown ability of an RNAi reagent (Stealth RNAi™ siRNA, siRNA, dicer pools, or shRNA-containing plasmid). To identify the most effective knockdown reagent for your target, simply clone your target gene into the pSCREEN-iT™ /lacZ-DEST Gateway® vector. Co-transfect the resulting construct with the knockdown reagent being tested. Expressed genes will be fused to β-galactosidase, enabling you to use β-galactosidase levels to measure the amount of RNAi-induced degradation of the target gene (Figure 1). The more effective any specific knockdown reagent is at mediating RNAi, the more degradation of the fusion mRNA will occur, lowering β-gal activity (Figure 2).

The BLOCK-iT™ RNAi Target Screening System includes the pSCREEN-iT™/lacZ-DEST Gateway® vector for generating a construct containing a lacZ/gene of interest fusion. This vector features:

attR sites immediately following the lacZ ORF for fast and efficient recombination with any attL-flanked Gateway® entry vector to generate the lacZ/target gene fusions
•The CMV promoter for strong expression of the lacZ/target gene fusion, leading to increased assay sensitivity and better discrimination between knockdown reagents of varied effectiveness
•Positional cloning to fuse a gene or fragment to the 3´ end of the lacZ gene so that β-galactosidase will be expressed even if the target fragment being tested contains stop codons

In addition, the pSCREEN-iT™/lacZ-DEST vector is fully compatible with the Ultimate™ ORF Clone Collection. This makes a wide selection of potential target genes readily available for generating lacZ fusions.

BLOCK-iT™ Transfection Kit Invitrogen™

The BLOCK-iT™ Transfection Kit contains everything you need for successful high-efficiency transfection of siRNA into cells. Each kit contains Lipofectamine™ 2000 Transfection Reagent for efficient delivery in a broad range of cell lines and the BLOCK-iT™ Fluorescent Oligo for use as an indicator of transfection efficiency. The BLOCK-iT™ Fluorescent Oligo provides:

–A strong, easily detectable fluorescein signal indicating transfection efficiency

–Chemical modifications for a clearer, more persistent cellular signal than labeled siRNA
–Proven correlation of transfection efficiency with Stealth RNAi™ siRNA and traditional siRNA molecules

Silencer™ siRNA Starter Kit Invitrogen™

Perfect for researchers new to RNAi, this Ambion® kit contains all of the reagents, protocols, and instructions necessary to get underway with siRNA transfection and ensure successful gene knockdown experiments.

• The perfect choice for those beginning siRNA experiments
• Includes transfection agent, siRNAs, and three different systems for assessing knockdown
• For use with human, mouse, and rat cells

The Silencer® siRNA Starter Kit includes a high-quality, lipid-based transfection agent, siPORT™ NeoFX™, that effectively delivers siRNAs to a broad range of cell lines. It can be used for reverse transfection, a streamlined protocol that eliminates an entire day from the RNAi experimental procedure; it can also be used with standard transfection protocols. This reagent is not sensitive to serum and efficiently transfects low concentrations of siRNA. The kit also includes a well-characterized GAPDH siRNA, which has been shown to strongly down-regulate GADPH in human, mouse, and rat cells, as well as a nontargeting negative control siRNA, Silencer® Negative Control #1 siRNA. Because GAPDH is ubiquitously expressed and is easy to detect at both the mRNA and protein level, it is an ideal target for a positive control siRNA.

Three Detection Systems
The kit includes three different systems for monitoring GAPDH siRNA-induced knockdown. An anti-GAPDH antibody is provided to assess GAPDH protein levels by western blot or immunofluorescence. The KDalert™ GAPDH Assay is also provided to quickly measure GAPDH enzymatic activity. This assay can also be used to assess cell viability. Finally, RT-PCR primers are included, for real-time quantitative RT-PCR detection of GAPDH mRNA using SYBR® Green. These reagents can also be used to transfect miRNAs, such as the Ambion® Pre-miR™ miRNA Precursor Molecules.

MEGAscript™ RNAi Kit Invitrogen™

The Ambion® MEGAscript® RNAi Kit is for the synthesis of large mass amounts of dsRNA for RNAi experiments in nonmammalian systems. The MEGAscript RNAi Kit uses Ambion's patented high-yield transcription technology and includes reagents for transcription, nuclease digestion, and clean up. The kit includes sufficient reagents for 20 reactions.

Synthesize Large Amounts of dsRNA:
The ability to generate large amounts of clean, double-stranded RNA is essential for the success of RNAi experiments. The MEGAscript RNAi Kit allows RNAi researchers to synthesize 50 to 100 µg or more of double-stranded RNA in a single transcription reaction. The yield is typically 10–50 times the amount of RNA produced by conventional transcription reactions. DNA of either PCR or plasmid origin can be used as a template for the transcription reaction. The sense and antisense strands can be synthesized from a PCR-generated DNA template containing T7 RNA Polymerase promoters on both ends of the template, eliminating the need for a separate annealing step.

Silencer™ siRNA Construction Kit with manual Invitrogen™

The Ambion® Silencer® siRNA Construction Kit is for the synthesis of siRNAs by in vitro transcription of DNA templates and costs less per siRNA than chemical synthesis. The kit includes sufficient reagents to synthesize 15 siRNAs.
• Cut siRNA preparation time—generate up to 15 siRNAs in less than 24 hours
• Get hundreds of transfections per reaction

Silencer™ siRNA Transfection II Kit Invitrogen™

This Ambion® kit is for the optimization of transfection protocols for specific cell types. It includes 0.4 mL of each of two transfection agents; siPORT™ Amine Transfection Agent and siPORT™ NeoFX™ Transfection Agent along with control siRNAs.
• Broad cell line compatibility-kit includes siPORT™ Amine and siPORT™ NeoFX™ transfection agents
• Faster protocol optimization-kit includes ready-to-transfect GAPDH and negative siRNA controls The choice of transfection agent is critical for gene silencing experiments. Every transfection agent has a different delivery efficiency when used with different cell types, and the overall transfection efficiency and degree of gene silencing depend on the nature of the transfection agent/siRNA complex. Whether you are new to RNAi research or you want to expand your experiments to include different cell lines, the Silencer® siRNA Transfection II Kit enables you to optimize your transfection protocols for specific cell types. The Silencer® siRNA Transfection II Kit includes siPORT™ Amine Transfection Agent (a polyamine mixture) and the new siPORT™ NeoFX™ Transfection Agent (a cationic and neutral lipid mixture). These reagents support efficient, reproducible siRNA transfections in different cell types. Furthermore, both can be used to transfect cells during subculturing-saving a full day of valuable time-without increased cytotoxicity. In addition to containing a choice of transfection agents, this kit includes a well-characterized GAPDH synthetic siRNA that can be used to optimize transfection conditions. Results can also be verified at the protein level with GAPDH-specific antibodies. A scrambled, negative control siRNA is also provided to confirm the specificity of the siRNA of interest and to demonstrate the absence of nonspecific, toxic effects.

MEGAscript™ RNAi Kit with Manual Invitrogen™

The Ambion® MEGAscript® RNAi Kit is for the synthesis of large mass amounts of dsRNA for RNAi experiments in nonmammalian systems. The MEGAscript RNAi Kit uses Ambion's patented high-yield transcription technology and includes reagents for transcription, nuclease digestion, and clean up. The kit includes sufficient reagents for 20 reactions.

Synthesize Large Amounts of dsRNA:
The ability to generate large amounts of clean, double-stranded RNA is essential for the success of RNAi experiments. The MEGAscript RNAi Kit allows RNAi researchers to synthesize 50 to 100 µg or more of double-stranded RNA in a single transcription reaction. The yield is typically 10–50 times the amount of RNA produced by conventional transcription reactions. DNA of either PCR or plasmid origin can be used as a template for the transcription reaction. The sense and antisense strands can be synthesized from a PCR-generated DNA template containing T7 RNA Polymerase promoters on both ends of the template, eliminating the need for a separate annealing step.
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