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RNAi in Non-Mammalian Cultured Cells

promoters results in the annealing of ssRNAs in the reaction tube.

Figure 1. MEGAscript RNAi Kit Procedure.


To ensure that any effects seen in an RNAi experiment are due to the dsRNA itself, it is imperative that the RNA be purified from any contaminants carried over from the transcription reaction. Unlike less complete competing products, the MEGAscript RNAi Kit provides all the reagents necessary to eliminate DNA template, single-stranded RNA (ssRNA), proteins and free nucleotides. After the dsRNA is synthesized, it is digested with DNase I and ssRNA specific RNase (included) to degrade the DNA template and any residual ssRNA, respectively. The dsRNA is then purified away from proteins, free nucleotides, and nucleic acid degradation products using a quick glass fiber filter-based procedure. The kit also comes with a comprehensive Instruction Manual and a control template to assess transcriptional efficiency. Up to 2 mg of dsRNA can be produced from a single kit.


Analysis of the RNAi Effect
Ambion has developed a complete line of products and a unique web resource for the design, execution and analysis of RNAi experiments (see www.ambion.com/RNAi). Our kits and reagents have been developed and are supported by scientists at Ambion who are actively performing silencing experiments of their own (Jarvis and Ford, 2001; Brown et al., 2002; Byron et al., 2002). Here we have tested and optimized various paramet
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