RNA Analyzer³ Tutorial

Welcome to RNA Analyzer³ tutorial! This brief guide will walk you through the process of analyzing your RNA/DNA sequences using our comprehensive web application.

What is RNA Analyzer?

RNA Analyzer is a web-based tool that allows you to search your RNA and/or DNA sequences for several known RNA structures and retrieve general information about your sequences. Check out out help page to know more about the scans it offers.

1. Input

RNA Analyzer accepts RNA and/or DNA sequences in fasta format. Users can input the sequences in two ways:

Option 1: Direct Sequence Entry

Option 2: FASTA File Upload

3. Selecting Motif Scans

RNA Analyzer offers several types of motif scans. By default, all motif scans are enabled, but you can chose which ones to perform:

Available Motif Scans

Iron Response Elements (IRE): Identifies iron-responsive elements that regulate iron metabolism in cells.

RNA Motif Scan: Searches for various known RNA structural motifs.

Protein Binding Motif Scan: Finds RNA-binding protein recognition sites.

Trans-splicing Structure: Detects trans-splicing signals and structures.

tRNA Scan: Detects transfer RNA sequences and secondary structures.

miRNA Scan: Identifies microRNA sequences and precursors.

4. Advanced Options

Coding Prediction with Augustus

You can optionally run gene prediction on your sequence using the Augustus software:

Available species models:

• Human (Homo sapiens)
• C. elegans (Caenorhabditis elegans)
• E. coli (Escherichia coli K12)
• A. thaliana (Arabidopsis thaliana)
• S. cerevisiae (Saccharomyces cerevisiae)
• D. melanogaster (Drosophila melanogaster)
• D. rerio (Danio rerio/Zebrafish)

miRNA Target Scan

5. Interpreting Results

Your results are organized into expandable sections with a tabular styled output for easier interpretation. Click on any section header to view its contents. Each section contains different types of analysis:

Structural Information Section

This section provides overall sequence statistics and structural analysis:

• Length: Total sequence length in nucleotides
• Energy: Predicted folding energy (kcal/mol) - more negative values indicate more stable structures
• Stems: Number of identified stem-loop structures
• Hairpins: Number of terminal hairpin structures detected
• Comment: SimpleiInterpretation of structural complexity
• Stem GG pairs: Special GG base pairs that may indicate specific structural motifs

Individual Motif Scan Results

Trans-Splicing Analysis: Searches for trans-splicing signals in Schistosoma and C. elegans patterns

Iron Response Elements (IRE): Identifies iron-responsive regulatory elements

Au-rich regions (ARE): Finds AU-rich elements that affect mRNA stability, showing sequence, position, and number of mismatches

Catalytic RNA sites: Detects snRNP motifs and putative Sm-sites involved in splicing. Quality scores (+ or ++) indicate prediction confidence

RNA Motif Scan: Uses Rfam database matching to identify known RNA families. Each hit includes:

• Rfam family ID with direct links to database
• Start/end positions and E-values
• Interactive structure viewer: Click "View Structure" to view RNA structure visualization

RNA Type Scans:

tRNA Scan: Identifies transfer RNA sequences using tRNAscan-SE

RNA Binding Protein Motifs: Finds binding sites for specific proteins like PTBP1, NOVA2, etc. Shows:

• Protein name and motif type (e.g., RRMx4, KHx3)
• Position, score, and p-value
• Exact matched sequence

miRNA Scan: Detects microRNA precursors with links to miRBase database entries

Optional Analysis Results (if enabled)

Gene Prediction Analysis: When Augustus is enabled, shows predicted exons with:

• Exon boundaries and types (coding vs. noncoding)
• Transcript structure information
• Predicted protein sequence

UTR Prediction: Identifies 5' and 3' untranslated regions showing:

• UTR boundaries and length
• Secondary structure information (stems, energy)
• Regulatory motifs (Shine-Dalgarno, polyA signals)

miRNA Target Prediction: If enabled, shows potential miRNA binding sites in 3' UTRs with binding energy and score

Interactive RNA Structure Visualization

We also provide a comprehensive RNA structure visualization at the bottom:

Color Legend for Structure Visualization

• ■ Red: General motifs
• ■ Light Blue: UTRs (untranslated regions)
• ■ Green: Exons (coding sequences)
• ■ Orange: tRNA structures
• ■ Yellow: miRNA sequences
• ■ Purple: Sm-sites and snRNP motifs
• ■ Cyan: Trans-splicing elements
• ■ Lime: PolyA signals and tails
• ■ Pink: Protein binding sites

Feature Summary Table

At the bottom, you'll find a comprehensive table summarizing features with their coordinates. This table is useful for:

• Getting precise coordinates for downstream analysis
• Seeing all features at a glance
• Identifying overlapping or nearby features

6. Troubleshooting

Common Issues