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Tom Tuschl s siRNA Design Commandments

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  • siRNA duplexes should be composed of 21-nucleotide sense and antisense strands, paired to have a 2-nt 3’ overhang (a TT overhang tends to work well).
  • A particularly effective target motif is NAR(N17)YNN, where N is any nucleotide, R is an A or a G, and Y is a C or a U.
  • Target region should be selected from a cDNA sequence beginning 50 to 100 nucleotides downstream of the start codon.
  • Target cDNA sequence should have around 50 percent GC content.
  • Avoid more than three consecutive identical nucleotides.
  • Secondary structure of the target mRNA does not appear to affect silencing.
  • Selected siRNA sequences should be Blasted against EST libraries to ensure that only one gene is targeted.

Source: The siRNA User Guide: http://www.mpibpc.gwdg.de/abteilungen/100/105/sirna.html

 

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