@bgicli/bgicli 2.1.1 → 2.2.0

package/data/skills/tooluniverse-multi-omics-integration/SKILL.md

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+---
+name: tooluniverse-multi-omics-integration
+description: Integrate and analyze multiple omics datasets (transcriptomics, proteomics, epigenomics, genomics, metabolomics) for systems biology and precision medicine. Performs cross-omics correlation, multi-omics clustering (MOFA+, NMF), pathway-level integration, and sample matching. Coordinates ToolUniverse skills for expression data (RNA-seq), epigenomics (methylation, ChIP-seq), variants (SNVs, CNVs), protein interactions, and pathway enrichment. Use when analyzing multi-omics datasets, performing integrative analysis, discovering multi-omics biomarkers, studying disease mechanisms across molecular layers, or conducting systems biology research that requires coordinated analysis of transcriptome, genome, epigenome, proteome, and metabolome data.
+---
+
+# Multi-Omics Integration
+
+Coordinate and integrate multiple omics datasets for comprehensive systems biology analysis. This skill orchestrates specialized ToolUniverse skills to perform cross-omics correlation, multi-omics clustering, pathway-level integration, and unified interpretation across molecular layers.
+
+## When to Use This Skill
+
+**Triggers**:
+- User has multiple omics datasets (RNA-seq + proteomics, methylation + expression, etc.)
+- Requests for integrative multi-omics analysis
+- Cross-omics correlation queries (e.g., "How does methylation affect expression?")
+- Multi-omics biomarker discovery
+- Systems biology questions requiring multiple molecular layers
+- Precision medicine applications with multi-omics patient data
+- Questions about molecular mechanisms across omics types
+
+**Example Questions This Skill Solves**:
+1. "Integrate RNA-seq and proteomics data to find genes with concordant changes"
+2. "How does promoter methylation correlate with gene expression?"
+3. "Perform multi-omics clustering to identify patient subtypes"
+4. "Which pathways are dysregulated across transcriptome, proteome, and metabolome?"
+5. "Find multi-omics biomarkers for disease classification"
+6. "Correlate CNV with gene expression to identify dosage effects"
+7. "Integrate GWAS variants, eQTLs, and expression data"
+8. "Perform MOFA+ analysis on multi-omics cancer data"
+
+---
+
+## Core Capabilities
+
+| Capability | Description |
+|-----------|-------------|
+| **Data Integration** | Match samples across omics, handle missing data, normalize scales |
+| **Cross-Omics Correlation** | Correlate features across molecular layers (gene expression vs protein, methylation vs expression) |
+| **Multi-Omics Clustering** | MOFA+, NMF, joint clustering to identify omics-driven subtypes |
+| **Pathway Integration** | Combine omics evidence at pathway level for unified biological interpretation |
+| **Biomarker Discovery** | Identify multi-omics signatures with improved predictive power |
+| **Skill Coordination** | Orchestrate RNA-seq, epigenomics, variant-analysis, protein-interactions, gene-enrichment skills |
+| **Visualization** | Circos plots, integrated heatmaps, network visualizations |
+| **Reporting** | Unified multi-omics reports with cross-layer insights |
+
+---
+
+## Workflow Overview
+
+```
+Input: Multiple Omics Datasets
+    |
+    v
+Phase 1: Data Loading & QC
+    |-- Load RNA-seq (expression matrix)
+    |-- Load proteomics (protein abundance)
+    |-- Load methylation (beta values or M-values)
+    |-- Load variants (CNV, SNV from VCF)
+    |-- Load metabolomics (metabolite abundance)
+    |-- Quality control per omics type
+    |
+    v
+Phase 2: Sample Matching
+    |-- Match samples across omics by ID
+    |-- Identify common samples
+    |-- Handle batch effects
+    |-- Normalize sample identifiers
+    |
+    v
+Phase 3: Feature Mapping
+    |-- Map features to common identifier space (genes, proteins, metabolites)
+    |-- Link CpG sites to genes (promoter, gene body)
+    |-- Map variants to genes
+    |-- Create unified feature matrix
+    |
+    v
+Phase 4: Cross-Omics Correlation
+    |-- Gene expression vs protein abundance (translation efficiency)
+    |-- Promoter methylation vs expression (epigenetic regulation)
+    |-- CNV vs expression (dosage effect)
+    |-- eQTL variants vs expression (genetic regulation)
+    |-- Metabolite vs enzyme expression (metabolic flux)
+    |
+    v
+Phase 5: Multi-Omics Clustering
+    |-- MOFA+ (Multi-Omics Factor Analysis) for latent factors
+    |-- NMF (Non-negative Matrix Factorization) for patient subtypes
+    |-- Joint clustering across omics
+    |-- Identify omics-specific vs shared variation
+    |
+    v
+Phase 6: Pathway-Level Integration
+    |-- Aggregate omics to pathway level
+    |-- Score pathway dysregulation (combined evidence)
+    |-- Use ToolUniverse enrichment tools (Reactome, KEGG, GO)
+    |-- Identify driver pathways across omics
+    |
+    v
+Phase 7: Biomarker Discovery
+    |-- Feature selection across omics
+    |-- Multi-omics signatures for classification
+    |-- Cross-validation and performance
+    |-- Interpretation and biological validation
+    |
+    v
+Phase 8: Generate Integrated Report
+    |-- Summary statistics per omics
+    |-- Cross-omics correlation results
+    |-- Multi-omics clusters and subtypes
+    |-- Top dysregulated pathways
+    |-- Multi-omics biomarkers
+    |-- Biological interpretation
+```
+
+---
+
+## Phase Details
+
+### Phase 1: Data Loading & Quality Control
+
+**Objective**: Load multiple omics datasets and perform quality control.
+
+**Supported omics types**:
+- **Transcriptomics**: RNA-seq count matrices, microarray
+- **Proteomics**: Protein abundance (MS-based)
+- **Epigenomics**: Methylation (450K, EPIC arrays, WGBS), ChIP-seq peaks
+- **Genomics**: CNV, SNV, structural variants
+- **Metabolomics**: Metabolite abundance (targeted, untargeted)
+
+**Data formats**:
+- Expression: CSV/TSV matrices, HDF5, AnnData (.h5ad)
+- Proteomics: MaxQuant output, Spectronaut, DIA-NN
+- Methylation: IDAT files, beta value matrices
+- Variants: VCF, SEG files (CNV)
+- Metabolomics: Peak tables, identified metabolites
+
+**Quality control per omics**:
+```python
+# RNA-seq QC
+- Filter low-count genes (mean counts < threshold)
+- Normalize (TPM, FPKM, or DESeq2)
+- Log-transform for correlation
+
+# Proteomics QC
+- Filter proteins with high missing values
+- Impute missing values (minimum, KNN)
+- Normalize (median, quantile)
+
+# Methylation QC
+- Remove failed probes
+- Correct for batch effects (ComBat)
+- Filter cross-reactive probes
+
+# Variants QC
+- Use variant-analysis skill for VCF QC
+- CNV segmentation validation
+```
+
+### Phase 2: Sample Matching
+
+**Objective**: Identify common samples across omics datasets.
+
+**Sample ID harmonization**:
+```python
+def match_samples_across_omics(omics_data_dict):
+    """
+    Match samples across multiple omics datasets.
+
+    Parameters:
+    omics_data_dict: {
+        'rnaseq': DataFrame (genes x samples),
+        'proteomics': DataFrame (proteins x samples),
+        'methylation': DataFrame (CpGs x samples),
+        'cnv': DataFrame (genes x samples)
+    }
+
+    Returns:
+    - common_samples: List of sample IDs present in all omics
+    - matched_data: Dict of DataFrames with common samples only
+    """
+    # Extract sample IDs from each omics
+    sample_ids = {
+        omics_type: set(df.columns)
+        for omics_type, df in omics_data_dict.items()
+    }
+
+    # Find common samples (intersection)
+    common_samples = set.intersection(*sample_ids.values())
+
+    # Subset each omics to common samples
+    matched_data = {
+        omics_type: df[sorted(common_samples)]
+        for omics_type, df in omics_data_dict.items()
+    }
+
+    return sorted(common_samples), matched_data
+```
+
+**Handling missing omics**:
+- Pairwise integration if not all samples have all omics
+- Document sample availability matrix
+
+### Phase 3: Feature Mapping
+
+**Objective**: Map features from different omics to common gene-level identifiers.
+
+**Gene-centric integration**:
+```python
+# Map all features to genes
+feature_mapping = {
+    'rnaseq': 'gene_symbol',  # Already gene-level
+    'proteomics': 'gene_symbol',  # Map protein to gene
+    'methylation': 'gene_symbol',  # Map CpG to gene (promoter)
+    'cnv': 'gene_symbol',  # CNV regions to overlapping genes
+    'metabolomics': 'enzyme_gene'  # Metabolite to enzyme gene
+}
+```
+
+**CpG to gene mapping**:
+- **Promoter methylation**: CpGs within TSS ± 2kb
+- **Gene body methylation**: CpGs within gene boundaries
+- Average methylation per gene (weighted by probe coverage)
+
+**CNV to gene mapping**:
+- Use variant-analysis skill to identify genes in CNV regions
+- Calculate copy number per gene (log2 ratio)
+
+### Phase 4: Cross-Omics Correlation
+
+**Objective**: Correlate features across molecular layers to understand regulation.
+
+**Example analyses**:
+
+#### 4.1: Expression vs Protein (Translation Efficiency)
+
+```python
+def correlate_rna_protein(rnaseq_data, proteomics_data):
+    """
+    Correlate mRNA and protein levels for each gene.
+
+    Expected: Positive correlation (r ~ 0.4-0.6 typical)
+    Discordance indicates post-transcriptional regulation
+    """
+    # Find common genes
+    common_genes = set(rnaseq_data.index) & set(proteomics_data.index)
+
+    correlations = {}
+    for gene in common_genes:
+        rna = rnaseq_data.loc[gene]
+        protein = proteomics_data.loc[gene]
+
+        # Spearman correlation (robust to outliers)
+        r, p = spearmanr(rna, protein)
+        correlations[gene] = {'r': r, 'p': p}
+
+    # Identify discordant genes (low RNA-protein correlation)
+    discordant = {g: v for g, v in correlations.items() if abs(v['r']) < 0.2}
+
+    return correlations, discordant
+```
+
+#### 4.2: Methylation vs Expression (Epigenetic Regulation)
+
+```python
+def correlate_methylation_expression(methylation_data, rnaseq_data):
+    """
+    Correlate promoter methylation with gene expression.
+
+    Expected: Negative correlation (increased methylation → decreased expression)
+    """
+    # For each gene with promoter methylation
+    results = {}
+    for gene in methylation_data.index:
+        if gene in rnaseq_data.index:
+            meth = methylation_data.loc[gene]  # Average promoter beta
+            expr = rnaseq_data.loc[gene]
+
+            r, p = spearmanr(meth, expr)
+            results[gene] = {'r': r, 'p': p, 'direction': 'repressive' if r < 0 else 'activating'}
+
+    # Identify genes with strong methylation-expression anticorrelation
+    regulated = {g: v for g, v in results.items() if v['r'] < -0.5 and v['p'] < 0.01}
+
+    return results, regulated
+```
+
+#### 4.3: CNV vs Expression (Dosage Effect)
+
+```python
+def correlate_cnv_expression(cnv_data, rnaseq_data):
+    """
+    Correlate copy number with gene expression.
+
+    Expected: Positive correlation (gene dosage effect)
+    """
+    results = {}
+    for gene in cnv_data.index:
+        if gene in rnaseq_data.index:
+            cnv = cnv_data.loc[gene]  # log2 ratio
+            expr = rnaseq_data.loc[gene]
+
+            r, p = pearsonr(cnv, expr)
+            results[gene] = {'r': r, 'p': p}
+
+    # Genes with dosage effect (CNV drives expression)
+    dosage_genes = {g: v for g, v in results.items() if v['r'] > 0.5 and v['p'] < 0.01}
+
+    return results, dosage_genes
+```
+
+### Phase 5: Multi-Omics Clustering
+
+**Objective**: Identify patient subtypes using integrated omics data.
+
+**Method 1: MOFA+ (Multi-Omics Factor Analysis)**
+
+MOFA+ identifies latent factors that explain variation across omics.
+
+```python
+# Conceptual workflow (uses R's MOFA2 package or Python implementation)
+# 1. Prepare multi-omics data as list of matrices
+# 2. Run MOFA+ to identify factors
+# 3. Inspect factor variance explained per omics
+# 4. Cluster samples based on factor scores
+
+# Example interpretation:
+# Factor 1: Explains 40% variance in RNA-seq, 30% in proteomics → Cell proliferation
+# Factor 2: Explains 50% variance in methylation → Epigenetic subtype
+# Factor 3: Explains 20% variance in CNV → Genomic instability
+```
+
+**Method 2: Joint NMF (Non-negative Matrix Factorization)**
+
+Decompose multi-omics matrices into shared latent components.
+
+```python
+def joint_nmf_clustering(omics_data_dict, n_clusters=3):
+    """
+    Perform joint NMF across omics for clustering.
+
+    Returns patient cluster assignments based on shared factors.
+    """
+    # Concatenate omics matrices (after normalization)
+    combined_matrix = np.vstack([
+        omics_data_dict['rnaseq'].values,
+        omics_data_dict['proteomics'].values,
+        omics_data_dict['methylation'].values
+    ])
+
+    # Run NMF
+    from sklearn.decomposition import NMF
+    model = NMF(n_components=n_clusters, init='nndsvd', random_state=42)
+    W = model.fit_transform(combined_matrix)  # Feature loadings
+    H = model.components_  # Sample coefficients
+
+    # Cluster samples based on H (components)
+    from sklearn.cluster import KMeans
+    clusters = KMeans(n_clusters=n_clusters).fit_predict(H.T)
+
+    return clusters, W, H
+```
+
+**Method 3: Similarity Network Fusion (SNF)**
+
+Integrate omics through patient similarity networks.
+
+### Phase 6: Pathway-Level Integration
+
+**Objective**: Aggregate multi-omics evidence at the pathway level.
+
+**Approach**: Score pathway dysregulation using combined evidence from multiple omics.
+
+```python
+def integrate_pathway_evidence(omics_results, pathway_genes):
+    """
+    Score pathway dysregulation across omics.
+
+    omics_results: {
+        'rnaseq': {'gene': fold_change},
+        'proteomics': {'gene': fold_change},
+        'methylation': {'gene': methylation_diff},
+        'cnv': {'gene': copy_number}
+    }
+
+    pathway_genes: List of genes in pathway
+    """
+    # For each gene in pathway
+    pathway_scores = []
+    for gene in pathway_genes:
+        gene_score = 0
+        evidence_count = 0
+
+        # RNA-seq evidence
+        if gene in omics_results['rnaseq']:
+            gene_score += abs(omics_results['rnaseq'][gene])
+            evidence_count += 1
+
+        # Proteomics evidence
+        if gene in omics_results['proteomics']:
+            gene_score += abs(omics_results['proteomics'][gene])
+            evidence_count += 1
+
+        # Methylation evidence (negative correlation)
+        if gene in omics_results['methylation']:
+            gene_score += abs(omics_results['methylation'][gene])
+            evidence_count += 1
+
+        # CNV evidence
+        if gene in omics_results['cnv']:
+            gene_score += abs(omics_results['cnv'][gene])
+            evidence_count += 1
+
+        if evidence_count > 0:
+            pathway_scores.append(gene_score / evidence_count)
+
+    # Aggregate pathway score (mean of gene scores)
+    pathway_score = np.mean(pathway_scores) if pathway_scores else 0
+
+    return {
+        'pathway_score': pathway_score,
+        'n_genes_with_evidence': len(pathway_scores),
+        'n_omics_types': evidence_count
+    }
+```
+
+**Use ToolUniverse enrichment tools**:
+```python
+# Get pathways for gene set
+from tooluniverse import ToolUniverse
+tu = ToolUniverse()
+
+# Enrichment for genes dysregulated in ANY omics
+all_dysregulated_genes = set()
+all_dysregulated_genes.update(rnaseq_degs)
+all_dysregulated_genes.update(diff_proteins)
+all_dysregulated_genes.update(methylation_dmgs)
+
+# Run enrichment
+enrichment = tu.run_one_function({
+    "name": "enrichr_enrich",
+    "arguments": {
+        "gene_list": ",".join(all_dysregulated_genes),
+        "library": "KEGG_2021_Human"
+    }
+})
+
+# Score each pathway with multi-omics evidence
+for pathway in enrichment['data']['results']:
+    pathway_genes = pathway['genes']
+    pathway['multi_omics_score'] = integrate_pathway_evidence(
+        omics_results, pathway_genes
+    )
+```
+
+### Phase 7: Biomarker Discovery
+
+**Objective**: Identify multi-omics signatures for disease classification.
+
+**Feature selection across omics**:
+```python
+def select_multiomics_features(X_dict, y, n_features=50):
+    """
+    Select top features across omics for classification.
+
+    X_dict: {
+        'rnaseq': DataFrame (samples x genes),
+        'proteomics': DataFrame (samples x proteins),
+        'methylation': DataFrame (samples x CpGs)
+    }
+    y: Target labels (disease vs control)
+
+    Returns: Selected features per omics
+    """
+    from sklearn.feature_selection import SelectKBest, f_classif
+
+    selected_features = {}
+    for omics_type, X in X_dict.items():
+        selector = SelectKBest(f_classif, k=min(n_features, X.shape[1]))
+        selector.fit(X, y)
+
+        # Get selected feature names
+        selected_idx = selector.get_support()
+        selected_features[omics_type] = X.columns[selected_idx].tolist()
+
+    return selected_features
+```
+
+**Multi-omics classification**:
+```python
+def multiomics_classification(X_dict, y, selected_features):
+    """
+    Train classifier using multi-omics features.
+    """
+    from sklearn.ensemble import RandomForestClassifier
+    from sklearn.model_selection import cross_val_score
+
+    # Concatenate selected features from each omics
+    X_combined = []
+    for omics_type, features in selected_features.items():
+        X_combined.append(X_dict[omics_type][features])
+
+    X_combined = pd.concat(X_combined, axis=1)
+
+    # Train classifier
+    clf = RandomForestClassifier(n_estimators=100, random_state=42)
+    scores = cross_val_score(clf, X_combined, y, cv=5, scoring='roc_auc')
+
+    return {
+        'mean_auc': scores.mean(),
+        'std_auc': scores.std(),
+        'n_features': X_combined.shape[1],
+        'features_per_omics': {k: len(v) for k, v in selected_features.items()}
+    }
+```
+
+### Phase 8: Integrated Reporting
+
+**Generate comprehensive multi-omics report**:
+
+```markdown
+# Multi-Omics Integration Report
+
+## Dataset Summary
+- **Omics Types**: RNA-seq, Proteomics, Methylation, CNV
+- **Common Samples**: 45 patients (30 disease, 15 control)
+- **Features**: 15,000 genes, 5,000 proteins, 450K CpGs, 20K CNV regions
+
+## Cross-Omics Correlation
+
+### RNA-Protein Correlation
+- **Overall correlation**: r = 0.52 (expected: 0.4-0.6)
+- **Highly correlated**: 3,245 genes (45%)
+- **Discordant genes**: 890 genes (post-transcriptional regulation)
+
+### Methylation-Expression
+- **Promoter methylation**: Anticorrelation r = -0.41
+- **Epigenetically regulated genes**: 1,256 genes (p < 0.01)
+- **Example**: BRCA1 promoter hypermethylation → 3-fold reduced expression
+
+### CNV-Expression Dosage Effect
+- **Genes with dosage effect**: 445 genes (r > 0.5, p < 0.01)
+- **Example**: MYC amplification (3 copies) → 2.8-fold increased expression
+
+## Multi-Omics Clustering
+
+### MOFA+ Analysis
+- **Factor 1** (25% variance): Cell cycle genes (RNA + protein)
+- **Factor 2** (18% variance): Immune signature (RNA + methylation)
+- **Factor 3** (15% variance): Metabolic reprogramming (RNA + metabolites)
+
+### Patient Subtypes
+- **Subtype 1** (n=18): High proliferation, MYC amplification
+- **Subtype 2** (n=15): Immune-enriched, hypomethylation
+- **Subtype 3** (n=12): Metabolic dysregulation, mitochondrial dysfunction
+
+## Pathway Integration
+
+### Top Dysregulated Pathways (Multi-Omics Score)
+1. **Cell Cycle** (score: 8.5) - RNA (↑), Protein (↑), CNV (amplification)
+2. **Immune Response** (score: 7.2) - RNA (↑), Methylation (hypo)
+3. **Glycolysis** (score: 6.8) - RNA (↑), Metabolites (↑)
+
+## Multi-Omics Biomarkers
+
+### Classification Performance
+- **AUC**: 0.92 ± 0.04 (5-fold CV)
+- **Features**: 50 total (20 RNA, 15 protein, 10 methylation, 5 CNV)
+- **Top biomarkers**:
+  - MYC expression (RNA)
+  - CDK1 protein abundance
+  - BRCA1 promoter methylation
+  - TP53 CNV status
+
+## Biological Interpretation
+
+The multi-omics analysis reveals three distinct disease subtypes driven by different molecular mechanisms:
+
+1. **Proliferative subtype**: Characterized by MYC amplification driving coordinated upregulation of cell cycle genes at both RNA and protein levels.
+
+2. **Immune subtype**: Hypomethylation of immune genes leading to increased expression and T-cell infiltration.
+
+3. **Metabolic subtype**: Shift from oxidative phosphorylation to glycolysis, with concordant changes in enzyme expression and metabolite levels.
+
+These subtypes may respond differently to targeted therapies.
+```
+
+---
+
+## ToolUniverse Skills Coordination
+
+This skill orchestrates multiple specialized skills:
+
+| Skill | Used For | Phase |
+|-------|----------|-------|
+| `tooluniverse-rnaseq-deseq2` | Load and analyze RNA-seq data | Phase 1, 4 |
+| `tooluniverse-epigenomics` | Methylation analysis, ChIP-seq peaks | Phase 1, 4 |
+| `tooluniverse-variant-analysis` | CNV and SNV processing | Phase 1, 3, 4 |
+| `tooluniverse-protein-interactions` | Protein network context | Phase 6 |
+| `tooluniverse-gene-enrichment` | Pathway enrichment | Phase 6 |
+| `tooluniverse-expression-data-retrieval` | Public omics data retrieval | Phase 1 |
+| `tooluniverse-target-research` | Gene/protein annotation | Phase 3, 8 |
+
+---
+
+## Example Use Cases
+
+### Use Case 1: Cancer Multi-Omics
+
+**Question**: "Integrate TCGA breast cancer RNA-seq, proteomics, methylation, and CNV data"
+
+**Workflow**:
+1. Load 4 omics types for 500 patients
+2. Match samples (450 common across all omics)
+3. Correlate RNA-protein (identify translation-regulated genes)
+4. Correlate methylation-expression (find epigenetically silenced genes)
+5. Correlate CNV-expression (identify dosage-sensitive genes)
+6. Run MOFA+ to find latent factors
+7. Identify 4 subtypes with distinct multi-omics profiles
+8. Perform pathway enrichment per subtype
+9. Select multi-omics biomarkers (AUC=0.94)
+
+### Use Case 2: eQTL + Expression
+
+**Question**: "How do GWAS variants affect gene expression through methylation?"
+
+**Workflow**:
+1. Load genotype data (SNPs from GWAS)
+2. Load expression data (RNA-seq)
+3. Load methylation data (450K array)
+4. For each GWAS SNP:
+   - Test association with nearby gene expression (eQTL)
+   - Test association with nearby CpG methylation (meQTL)
+   - Test CpG-gene correlation
+5. Identify SNP → methylation → expression regulatory chains
+6. Annotate with ToolUniverse (GWAS traits, gene function)
+
+### Use Case 3: Drug Response Multi-Omics
+
+**Question**: "Predict drug response using multi-omics profiles"
+
+**Workflow**:
+1. Load baseline multi-omics (pre-treatment)
+2. Load drug response data (IC50 or clinical response)
+3. Correlate each omics with response
+4. Select multi-omics features predictive of response
+5. Train multi-omics classifier
+6. Identify pathways associated with resistance/sensitivity
+7. Use ToolUniverse drug-repurposing skill for alternative options
+
+---
+
+## Advanced Analysis Patterns
+
+### Pattern 1: Omics-Driven Patient Stratification
+
+For precision medicine applications where patient stratification is goal.
+
+### Pattern 2: Multi-Omics Network Analysis
+
+Build integrated networks combining PPI, co-expression, regulatory interactions.
+
+### Pattern 3: Temporal Multi-Omics
+
+Longitudinal multi-omics data (time-series or treatment response).
+
+### Pattern 4: Spatial Multi-Omics
+
+Spatial transcriptomics + proteomics for tissue architecture.
+
+---
+
+## Quantified Minimums
+
+| Component | Requirement |
+|-----------|-------------|
+| Omics types | At least 2 omics datasets |
+| Common samples | At least 10 samples across omics |
+| Cross-correlation | Pearson/Spearman correlation computed |
+| Clustering | At least one method (MOFA+, NMF, or SNF) |
+| Pathway integration | Enrichment with multi-omics evidence scores |
+| Report | Summary, correlations, clusters, pathways, biomarkers |
+
+---
+
+## Limitations
+
+- **Sample size**: Multi-omics integration requires sufficient samples (n≥20 recommended)
+- **Missing data**: Some patients may not have all omics types
+- **Batch effects**: Different omics platforms/batches require careful normalization
+- **Computational**: Large multi-omics datasets may require significant memory/compute
+- **Interpretation**: Multi-omics results require domain expertise for biological validation
+
+---
+
+## References
+
+**Methods**:
+- MOFA+: https://doi.org/10.1186/s13059-020-02015-1
+- Similarity Network Fusion: https://doi.org/10.1038/nmeth.2810
+- Multi-omics review: https://doi.org/10.1038/s41576-019-0093-7
+
+**ToolUniverse Skills**:
+- See individual skill documentation for omics-specific methods