Kay Lab siRNA/shRNA/Oligo Optimal Design

shRNA (short hairpin RNAs) are artificial constructs that can be expressed endogenously. These expressed hairpins fold to form dsRNA, and Drosha and Dicer then act on these hairpins to create mature sequence, used by the RISC complex to target genes.

siRNA/shRNA/oligo Optimal Design considering off-target: Besides intended gene down-regulation, shRNAs can introduce off-target gene silencing. Our program design shRNA with higher on-target potency, and less off-target effect. Unlike many traditional design programs, which focus on seed complementarity, we systematically study the influence of different nucleotides in each shRNA guide strand position on unintended gene silencing, and finally assign a score matrix for determining the degree of off-target effect. When scanning a target sequence, This program calculates the score for every possible shRNA, and ranks them from highest to lowest score. Ranked top shRNAs are then selected for on-target potency, and less off-target effect.

Please cite: Gu S, Zhang Y, Jin L, Huang Y, Zhang F, Bassik MC, Kampmann M, Kay MA. Weakbase pairing in both seed and 3' regions reduces RNAi off-targets and enhances si/shRNA designs. Nucleic Acids Res. 2014 Sep 30. pii: gku854.

Step1: Providing sequence information (please provide either gene accession numbers or sequence in fasta format)

Accession Number


      accession number (maximum of 20, comma separated)



      Paste in a nucleotide sequence in fasta format


Step2: Choosing other parameters


Number of shRNAs per gene (between 1 and 20):

Length of guide strand:

Maximum length of homopolymers in each shRNA:

Output format:

Questions for siRNA/shRNA/oligo Optimal Design, please email yuezhang@stanford.edu or yue27.zhang@gmail.com for assistance.

Last modified 11/01/2014.