siRNA Knockdown Designer
siRNA candidate design following Tuschl rules with asymmetric 3′ overhangs, GC filtering and seed-region screening.
How it works
Candidate 21-mers are generated across the transcript and filtered on GC content, thermodynamic asymmetry at the duplex ends, absence of internal repeats, and avoidance of SNP positions. Asymmetry matters because it determines which strand RISC loads: a duplex with a less stable 5′ antisense end preferentially loads the intended guide.
Frequently asked questions
What makes an effective siRNA?
Effective siRNAs typically have 30–52% GC content, an A or U at the 5′ end of the antisense strand to favour correct RISC loading, no runs of four or more identical bases, and no strong internal secondary structure in the target region. Designing three or four independent siRNAs against the same transcript and confirming that phenotypes agree is the standard control against off-target effects.
What is the difference between siRNA and shRNA knockdown?
siRNA is a synthetic double-stranded RNA transfected directly into cells, giving rapid but transient knockdown lasting a few days. shRNA is encoded on a plasmid or viral vector, transcribed inside the cell and processed into siRNA, which allows stable long-term knockdown and selection of stable lines. siRNA suits short experiments; shRNA suits anything requiring sustained silencing.