Aim
This online training provides a practical, step-by-step approach to designing effective siRNA molecules for gene silencing. Participants will learn how siRNA works (RNA interference), how to select target regions, apply design rules, evaluate specificity and off-target risk, and prepare a clean, experiment-ready siRNA shortlist—supported by tool-based workflows and a mini capstone design project.
Program Objectives
- Understand RNAi Mechanism: Learn how siRNA mediates gene knockdown and what determines potency.
- Target Selection Skills: Identify the right transcript regions and avoid problematic sequence zones.
- Apply Design Rules: Use practical sequence guidelines (GC%, thermodynamics, motifs) to design candidates.
- Reduce Off-Target Effects: Learn seed-region risks, mismatch tolerance, and specificity checks.
- Tool-Based Workflow: Practice with common siRNA design and validation tools (concept + hands-on flow).
- Experiment Readiness: Produce a shortlist with annotations, controls, and validation plan.
Program Structure
Module 1: RNA Interference (RNAi) Essentials
- siRNA vs miRNA vs shRNA: what’s different and when each is used.
- How RISC loading and strand selection works (why asymmetry matters).
- What “good knockdown” depends on: accessibility, thermodynamics, and delivery.
- Common reasons siRNA fails in experiments (and how to prevent them).
Module 2: Choosing the Right Target (Gene, Transcript, Region)
- Gene IDs, isoforms, and transcript selection (avoiding wrong isoform targeting).
- Target regions: CDS vs UTRs—pros/cons and typical best practices.
- Avoiding problematic zones: SNP-rich regions, repeats, high secondary structure.
- Designing for species specificity (human/mouse/plant/cell-line context).
Module 3: Core siRNA Design Rules (Practical Guidelines)
- Length and structure: duplex basics (sense/antisense, 2-nt overhang concept).
- GC content guidance and why extremes reduce performance.
- Sequence motifs to prefer/avoid (e.g., internal repeats, long GC stretches).
- Thermodynamic asymmetry: improving guide strand selection.
Module 4: Off-Target Effects & Specificity Checking
- Seed-region effects: how miRNA-like off-targeting happens.
- Mismatch tolerance and unintended transcript binding.
- Basic specificity workflow: similarity search against transcriptome (conceptual).
- Strategies to minimize off-targets: design filtering + validation planning.
Module 5: Secondary Structure, Accessibility & Efficacy Prediction
- Why mRNA structure matters for siRNA access and RISC engagement.
- Accessibility and local folding concepts (practical interpretation).
- Scoring and ranking candidates: combining rule-based + predictive scoring.
- Choosing 3–5 candidates wisely instead of 20 random ones.
Module 6: Controls, Experimental Planning & Validation
- Essential controls: negative controls, positive controls, mock transfection.
- Concentration planning and dose-response thinking (avoiding toxicity artifacts).
- Validation methods: qPCR vs Western blot vs phenotype assays.
- Interpreting knockdown results and troubleshooting weak effects.
Module 7: Delivery Considerations (Overview for Practical Context)
- Transfection basics: lipid-based delivery and common pitfalls.
- Stability and degradation: why chemical modifications are used (overview).
- In vitro vs in vivo considerations: what changes in design priorities.
- Safety and ethics basics for gene silencing experiments.
Final Project
- Pick a gene (provided list or your own) and design an siRNA shortlist.
- Deliverables: 3–5 candidate sequences (or masked representation if needed), target region notes, off-target checks summary, and validation plan.
- Example projects: siRNA design for an oncogene, inflammatory marker gene, viral entry protein, or pathway regulator.
Participant Eligibility
- UG/PG/PhD students in Biotechnology, Genetics, Molecular Biology, Bioinformatics, or related fields
- Researchers planning gene knockdown experiments in cell culture
- Life science professionals entering RNA therapeutics and functional genomics
- Basic understanding of DNA/RNA and gene expression is recommended
Program Outcomes
- Design Confidence: Ability to design and shortlist potent siRNAs using systematic rules.
- Specificity Awareness: Ability to anticipate and reduce off-target risks.
- Experiment Readiness: Produce a clean plan with controls and validation strategy.
- Troubleshooting Skills: Understand why knockdown fails and how to fix common issues.
- Portfolio Deliverable: An siRNA design dossier (shortlist + rationale) you can showcase.
Program Deliverables
- Access to e-LMS: Full access to training content and reference resources.
- Design Templates: siRNA screening checklist, off-target review sheet, validation plan template.
- Hands-on Exercises: Guided practice tasks for target selection and candidate ranking.
- Project Guidance: Mentor support for designing and refining your final shortlist.
- Final Assessment: Certification after assignments + final project submission.
- e-Certification and e-Marksheet: Digital credentials provided upon successful completion.
Future Career Prospects
- RNA Therapeutics Research Intern / Associate
- Functional Genomics Research Assistant
- Gene Silencing & Molecular Biology Associate
- Bioinformatics Support (RNAi / Target Design)
- Preclinical Research Associate (RNA-based pipelines)
Job Opportunities
- Biotech & Pharma: RNA therapeutics, target validation, and discovery research teams.
- Academic Research Labs: Functional genomics and gene regulation studies.
- CROs: Preclinical assay and molecular biology service teams.
- Healthtech Startups: RNA tool development and computational biology support roles.
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