siDirect

siDirect predicts effective small interfering RNA (siRNA) sequences for mammalian RNA interference (RNAi) by applying empirically derived sequence-design rules to optimize gene silencing.

Key Features:

• Sequence Input Flexibility: Accepts arbitrary nucleotide sequences and generates a list of candidate siRNAs.
• Guideline-Based Design: Applies specific, empirically validated siRNA design rules derived from extensive sequence–efficacy studies.
• Antisense Strand Requirements: Requires an A/U nucleotide at the 5' end of the antisense strand and a minimum of five A/U residues within the first one-third of the antisense strand.
• Sense Strand Requirement: Prefers a G/C nucleotide at the 5' end of the sense strand.
• GC Content Regulation: Excludes candidates containing any GC stretch longer than nine nucleotides.
• Cross-Hybridization Avoidance: Identifies and excludes siRNA candidates with potential off-target hybridization, addressing limitations encountered with traditional BLAST searches.
• Broad Applicability: Design rules are validated primarily for mammalian cells and are applicable to DNA-based RNAi in mammalian cells and in ovo RNAi in chick embryos, while sequence preferences are less discernible in Drosophila in vivo RNAi.

Scientific Applications:

• Functional Genomics: Design of siRNAs for systematic loss-of-function studies and gene function characterization in mammalian systems.
• Therapeutic Gene Silencing: Generation of siRNA candidates for research into gene-silencing therapeutic strategies in mammalian models.
• In ovo and DNA-based RNAi Studies: Support for siRNA design in in ovo RNAi experiments in chick embryos and DNA-based RNAi approaches in mammalian cells.

Methodology:

Processes input nucleotide sequences to generate candidate siRNAs, applies empirically derived sequence filters (antisense 5' A/U; ≥5 A/U in first one-third of antisense; sense 5' G/C; no GC stretch >9 nt), and excludes candidates with potential cross-hybridization to minimize off-target effects relative to BLAST-based checks.