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Design, Manufacture, and Assay of the Efficacy of siRNAs for Gene Silencing
Small interfering RNAs (siRNAs) have been widely exploited for nucleotide-sequence-specific posttranscriptional gene silencing, as a tool to investigate gene function in eukaryotes, and they hold promise as potential therapeutic agents. Conventionally designed siRNAs are 21-mers with symmetric 2-nt 3′ overhangs that mimic intermediates (microRNAs or miRNAs) of the natural processing of longer dsRNA (double-stranded RNA). siRNAs are sequences with full c omplementarity to their target mRNA and can be generated by either chemical synthesis or processing of shRNAs (short hairpin RNAs) transcribed from DNA vectors. To minimize off-target effects, any homology to nontarget mRNA can be verified using the expressed sequence tag (EST) database for the relevant organism. Here, we provide a practical guide and an overview to the design and selection of effective and specifc siRNAs.
- Production and Purification of Recombinant Adeno-Associated Vectors
- DNase i footprinting using PCR-generated end-labeled DNA probes
- Progress in the Therapeutic Applications of siRNAs Against HIV-1
- Separation, Digestion, and Cloning of Intact Parasite Chromosomes Embedded in Agarose
- Inducible Expression Cassettes in Yeast: GAL4
- Analysis of MicroRNA Expression Using Machine Learning
- Retroviral Infection
- MicroRNA Biogenesis: Isolation and Characterization of the Microprocessor Complex
- Transgenic Production Benchmarks
- Semi-Quantitative Detection of Hepatitis C Virus RNA by Real-Time RT-PCR
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