siRNA Design and RNA Secondary Structure Prediction

This internship is designed to introduce learners to the fundamentals and applications of RNA-based therapeutics, focusing on siRNA (small interfering RNA) design and RNA secondary structure prediction. Participants will learn about RNA interference (RNAi), target gene silencing, and how computational tools are used to predict RNA folding, thermodynamics, and stability.

Learning Objectives

By the end of the internship, participants will:

• Understand the principles of RNA interference and siRNA-mediated gene silencing

• Design effective siRNA sequences against target genes

• Predict RNA secondary structures and folding patterns

• Analyze RNA thermodynamic stability and hairpin loop formations

• Utilize online and open-source tools for RNA simulation and visualization

Project Structure

Introduction to RNAi and siRNA

• Mechanism of RNA interference

• siRNA vs miRNA vs shRNA

• Applications in therapeutics and gene regulation

Guidelines for siRNA Design

• siRNA structure and functional rules

• Off-target effects and GC content considerations

• Target site selection in mRNA

siRNA Design Tools

• Tools: siDirect, BLOCK-iT, DSIR, Whitehead siRNA Selection

• Hands-on: Designing siRNAs for target genes (e.g., BCL2, EGFR)

Advanced Filtering for siRNA

• Thermodynamics, target accessibility, specificity scoring

• Avoiding immune response triggers

Introduction to RNA Secondary Structures

• RNA folding, base pairing, stem-loops, hairpins

• RNA structure formats: dot-bracket, CT files

RNA Structure Prediction Tools

• Tools: RNAfold (ViennaRNA), Mfold, RNAstructure

• Hands-on: Predicting folding patterns and visualizing base-pair probabilities

Thermodynamic Analysis

• Free energy (ΔG) interpretation

• Structure ensemble generation

• Pseudoknot prediction (optional)

Visualization and Annotation

• Tools: Forna, VARNA, RNAstructure GUI

• Annotating loops, stems, bulges, and base-pair probabilities

Research Case Study

• Case Study: siRNA design against viral or cancer target

• Discussion of real-world RNAi-based therapies

Final Project Presentation

• Students design siRNA + RNA folding map for a selected gene

• Presentation of design rationale, structure prediction, and report submission

Tools & Resources

• siDirect, BLOCK-iT, DSIR – siRNA design

• RNAfold (ViennaRNA), Mfold, RNAstructure – RNA secondary structure prediction

• Forna, VARNA – RNA structure visualization

• BLAST, Ensembl, NCBI – Target gene selection

Assignments & Capstone Project

• Design siRNA sequences for 2 real genes

• Predict and compare RNA secondary structures

• Submit a report with sequences, structure visuals, and design evaluation

Certification

Participants receive:

• Certificate of Completion

• Project Evaluation Letter for academic or research use

Target Audience

• Graduate/PhD students in bioinformatics, molecular biology, or biotechnology

• Researchers in genetic engineering or RNA therapeutics

• Professionals interested in gene silencing or computational RNA biology