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Quantifies biodiversity from species abundance/incidence tables using Hill numbers (iNEXT) with coverage-based rarefaction-extrapolation (Chao & Jost 2012), asymptotic richness via Chao1/ACE/jackknife as a lower bound, Baselga turnover/nestedness partition with the Podani alternative as sensitivity check, mandatory Hellinger transformation before ordination (Legendre & Gallagher 2001), Faith PD and SES_MPD/SES_MNTD with explicit null-model choice, and Maire 2015 functional-diversity dimensionality optimization. Use when comparing diversity across sites with unequal sampling effort, picking the right richness estimator for singleton-heavy amplicon data, partitioning beta diversity into turnover vs nestedness, reporting Hill-number effective species counts rather than raw entropies, computing SES_MPD with explicit null-model justification, or deciding whether to apply standard metrics to compositional amplicon data. Not for clinical 16S microbiome diversity (see microbiome/diversity-analysis).

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Visualize biological networks (PPI, gene-regulatory, co-expression, pathway) with layout algorithm choice (ForceAtlas2, Fruchterman-Reingold, Kamada-Kawai, hive plots), edge bundling, community-based coloring, and reproducible seeds using NetworkX, PyVis, igraph, and Cytoscape automation. Use when rendering biological networks for static publication, interactive HTML exploration, or Cytoscape-format export.

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Build publication-quality figures with matplotlib using the object-oriented Figure/Axes API, constrained_layout, rcParams customization, TrueType (Type-42) font embedding for journal submission, and CVD-safe palettes. Covers seaborn integration, common chart types, axis formatting, and the small gotchas that distinguish reproducible matplotlib from notebook scratch. Use when producing publication figures in Python — RNA-seq scatter, single-cell embeddings, generic biological plotting.

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Build local BLAST databases and run searches using NCBI BLAST+ command-line tools. Use when running >50 queries, building custom databases with -parse_seqids and -taxid, downloading prebuilt NCBI databases via update_blastdb.pl, choosing -task variants (megablast/dc-megablast/blastn/blastn-short), tuning soft/hard masking, scaling threads, or extracting hits with blastdbcmd. Encodes BLAST v5 vs v4 database format, taxonomy filtering, makeblastdb pitfalls.

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Bulk-query Ensembl BioMart (and other BioMart instances) for cross-database ID mapping, gene/transcript/exon coordinates, and ortholog tables. Use when batch-converting Ensembl IDs to other namespaces (HGNC, RefSeq, UniProt, Entrez), pulling gene coordinate tables for thousands of genes, building ortholog wide-tables across species, or replacing slow Ensembl REST loops with one-shot bulk export. Encodes BioMart's XML query format, R biomaRt vs Python pybiomart trade-off, mart-vs-dataset hierarchy, and the URL endpoint that's BioMart-specific (separate from rest.ensembl.org).

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Select colormaps and qualitative palettes for scientific figures using perceptual-uniformity, color-vision-deficiency safety, and luminance-monotonicity criteria. Covers Crameri scientific colormaps, viridis/cividis/magma, Okabe-Ito categorical, ColorBrewer, and the rainbow/jet critique. Use when choosing palettes for heatmaps, scatter, networks, or any encoding where color carries quantitative or categorical meaning.

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Analyzes single-cell pooled CRISPR screens (Perturb-seq, CROP-seq, Perturb-CITE-seq, ECCITE-seq, multiome) where each cell carries an sgRNA and a scRNA-seq / surface-protein / chromatin readout. Covers experimental design (direct-capture Perturb-seq Dixit 2016 vs CROP-seq 3'UTR-barcoded Datlinger 2017 vs ECCITE-seq vs Multiome), MOI for sgRNA assignment, escaper-cell filtering (Mixscape, Papalexi 2021), SCEPTRE NB GLM + permutation for low-MOI (Barry 2024 Genome Biol 25:124), the Pertpy framework, factor decomposition, genome-scale Perturb-seq (Replogle 2022 Cell, 2.5M cells), and per-perturbation single-cell DE. Use when running a single-cell CRISPR screen, choosing direct-capture vs CROP-seq architecture, filtering escaper cells, performing single-cell DE, integrating Perturb-seq with pathway analysis, scaling to GW CRISPRi via Replogle protocol, or analyzing multi-omics screens.

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Predict RBP binding from RNA sequence using deep learning models (RBPNet sequence-to-signal, RNAProt RNN, GraphProt2 GCN with structure, DeepCLIP, DeepRiPe multi-modal CNN) for variant-effect prediction, in silico binding-site discovery, model interpretation, and transfer learning from CLIP and RBNS datasets. Use when computational prediction of RBP binding from sequence is needed, evaluating variant effects on binding without further wet-lab experiments, comparing model performance, or training a custom model on ENCODE eCLIP data.

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Model gene-family birth-death dynamics across a species tree using CAFE5 (Mendes et al 2020 Bioinformatics 36:5516 gamma-distributed rate categories), CAFE5-error (annotation-error-aware), Count (Csurös 2010 ancestral state reconstruction), BadiRate (Librado 2012 likelihood + parsimony), DupliPHY-Family, and ALE/AleRax (for per-family DTL; see [[gene-tree-species-tree-reconciliation]]). Test lineage-specific gene-family expansions and contractions, distinguish biological dynamics from annotation artifacts, account for assembly fragmentation, identify functional enrichment in expanded / contracted families. Use when correlating gene-family changes with phenotype evolution, ranking lineages by adaptive gene-family-rate shifts, post-WGD dosage-balance analysis, or building Birth-death models from OrthoFinder presence/absence matrices.

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Project gene annotations across genomes using TOGA (Kirilenko 2023 whole-genome-alignment chain-based projection with intactness classification), CESAR 2.0 (Sharma & Hiller 2017 codon-aware exon projection), LiftOff (Shumate & Salzberg 2020 reference-based annotation transfer), Liftover (UCSC), GeMoMa (Keilwagen 2019 evidence-based projection), and Comparative Annotation Toolkit (CAT). Use when transferring annotations from a well-annotated reference to query genome(s), classifying gene-loss vs gene-intact across many genomes at scale, building Zoonomia-style comparative annotations across hundreds of mammals or birds (Kirilenko 2023), detecting pseudogenization, projecting alternative isoforms, or selecting between WGA-anchored (TOGA) vs ortholog-based (LiftOff) annotation transfer strategies.

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Infer orthologous genes and gene families across species using OrthoFinder3 (HOG-based phylogenetic orthology), SonicParanoid2, Broccoli, ProteinOrtho, OMA / FastOMA hierarchical orthologous groups, eggNOG-mapper, JustOrthologs, and TOGA whole-genome-alignment orthology. Use when building single-copy ortholog sets for phylogenomics, classifying co-orthologs and in/out-paralogs after gene duplication, propagating functional annotation via orthology with awareness of the ortholog conjecture, distinguishing speciation from duplication via gene-tree species-tree reconciliation, computing Quest-for-Orthologs benchmark performance, or running synteny-aware ortholog detection in WGD-affected lineages.

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Detect positive (diversifying / episodic / pervasive) selection using codon dN/dS frameworks. Implements PAML codeml site models (M0/M1a/M2a/M7/M8/M8a), branch models, branch-site model A (Zhang 2005), and HyPhy methods (BUSTED, BUSTED-S, BUSTED-MH, BUSTED-PH, MEME, FEL, FUBAR, aBSREL, SLAC, RELAX, GARD, FUBAR-MH). Includes McDonald-Kreitman framework (asymptotic alpha, impMKT, polyDFE, DFE-alpha, GRAPES) for within-species + divergence inference, RERconverge for trait-correlated rate shifts, CSUBST for convergent substitution, and PhyloAcc for accelerated noncoding evolution. Use when testing adaptive evolution at codons, branches, or full gene; running GARD recombination pre-screen; controlling alignment-error and gBGC false positives; reconciling PAML vs HyPhy results; or performing genome-scale selection scans.

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Reconstruct ancestral states at internal phylogenetic nodes for sequences (PAML codeml, IQ-TREE --ancestral, GRASP, FastML), discrete traits (corHMM hidden-rate Markov, ape::ace, phytools::make.simmap stochastic mapping, BayesTraits), and continuous traits (phytools::fastAnc, geiger Brownian/OU, RPANDA). Use when designing constructs for ancestral protein resurrection, tracing trait evolution along a tree, performing stochastic character mapping, testing models of trait evolution (BM vs OU vs EB), inferring ancestral genome content via Dollo or DTL reconciliation, or quantifying ancestral-state uncertainty for downstream comparative analyses.

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Performs time-to-event analysis for clinical trials including Cox proportional hazards regression with PH diagnostics, restricted mean survival time (RMST) under non-PH, competing risks via Fine-Gray vs cause-specific Cox, weighted log-rank and MaxCombo for non-proportional hazards, recurrent events (Andersen-Gill, PWP, WLW), and interval-censored data. Use when analyzing time-to-event endpoints (OS, PFS, DOR, TTR, TTNT) in oncology or other clinical trials.

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Constructs and validates polygenic risk scores using LDpred2-auto, SBayesRC, MegaPRS, PRS-CS, PROSPER, MUSSEL, BridgePRS, JointPRS, PRSmix, or PGS Catalog Calculator with ancestry-aware reference panels (HapMap3, UKB-LD), Pejaver-style calibration, and PRS-RS reporting standards. Use when computing PRS for cohorts, applying Whiffin-style absolute-risk transformation, assessing cross-ancestry portability (Martin 2017 / Ding 2023 continuous ancestry), or auditing PRS manuscripts against the 22-item PRS-RS reviewer checklist.

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Performs subgroup and heterogeneous treatment effect (HTE) analyses for clinical trials. Covers Mantel-Haenszel pooling, Breslow-Day, interaction tests in regression, RERI for additive interaction, modern data-adaptive HTE methods (STEPP, SIDES, causal forests, X/R-learners), Bayesian shrinkage (Dixon-Simon, MAP, EXNEX), graphical multiplicity (Bretz-Maurer), and credibility frameworks (Sun BMJ, EMA 2019). Use when analyzing treatment effects across patient subgroups for regulatory submissions or precision-medicine claims.

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Designs adaptive clinical trials including group-sequential (O'Brien-Fleming, Pocock, Lan-DeMets spending), sample-size re-estimation (blinded Friede-Kieser, unblinded Cui-Hung-Wang, Mehta-Pocock promising zone), seamless Phase 2/3 with treatment-arm selection, population enrichment, and response-adaptive randomisation. Covers FDA 2019 Final Adaptive Designs Guidance, FDA 2022 Master Protocols, and ICH E20 Step 2b/3 draft (June 2025, NOT final). Use when planning interim analyses, sample-size re-estimation, or master/platform-trial designs.

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Performs logistic regression for clinical trial outcomes (binary, ordinal, multinomial) with marginal-vs-conditional estimand reporting per FDA 2023 covariate adjustment guidance, g-computation/standardisation for marginal effects, modified Poisson for RR, Brant test for proportional odds, Firth penalty for separation, and Hauck-Donner detection. Use when modeling binary or ordinal endpoints in confirmatory or exploratory clinical trials.

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Estimate causal effects of an exposure on an outcome from GWAS summary statistics using genetic instruments. Implements IVW (fixed/random), MR-Egger, weighted median/mode, MR-RAPS, CAUSE, GSMR-HEIDI, MR-PRESSO, MVMR, MR-Clust, LCV, and LHC-MR via TwoSampleMR, MendelianRandomization, MR-PRESSO, cause, and lhcMR. Use when testing causal direction between traits, evaluating drug-target effects via cis-pQTL/cis-eQTL, performing multivariable mediation MR, distinguishing causation from correlated horizontal pleiotropy, or producing STROBE-MR-compliant sensitivity batteries.

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Runs cis-pQTL Mendelian randomization for drug-target validation using UKB-PPP (Olink), deCODE (SomaScan), Fenland, INTERVAL, ARIC, and FinnGen-PPP proteomes plus colocalization triangulation, phenome-wide on-target adverse-effect scans, cross-platform Olink/SomaScan replication, and PAV (protein-altering variant) sensitivity. Use when nominating or de-risking a drug target from plasma-proteome GWAS, mimicking pharmacological inhibition via cis-pQTL instruments, separating shared-causal from LD-confounded signal under the Schmidt 2020 cis-MR framework, screening on-target adverse phenotypes pheWAS-style, or producing publication-grade STROBE-MR plus PP.H4 evidence for a target gene.

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Performs gene-level association from GWAS summary statistics via genetically predicted tissue expression using FUSION, PrediXcan, S-PrediXcan, S-MultiXcan, UTMOST, MOSTWAS, kTWAS, EpiXcan, TIGAR-V2, and probabilistic fine-mapping with FOCUS and MA-FOCUS. Use when running TWAS from GWAS sumstats, prioritising candidate causal genes from a GWAS lead locus, picking single-tissue vs cross-tissue models, identifying LD-induced TWAS false positives, choosing ancestry-matched prediction weights, fine-mapping co-regulated TWAS hits, or triangulating TWAS with cis-eQTL Mendelian randomization and colocalization to nominate a causal gene.

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Reads, writes, and converts molecular file formats (SMILES, InChI, SDF V2000/V3000, MOL2, PDB, MMTF) using RDKit and Open Babel with rigorous handling of aromaticity perception, stereochemistry, implicit/explicit hydrogens, kekulization, and salt/fragment separation. Use when loading chemical libraries, debugging parse failures, or preparing molecules for downstream standardization, descriptor calculation, or docking.

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Builds and applies 3D pharmacophore models using RDKit Pharm3D, the apo2ph4 receptor-based workflow (Heider et al 2022/2023 J Chem Inf Model 63:147-158), Pharmer / Pharmit (search), and PharmacoForge (diffusion-based generation, Flynn et al 2025 Front Bioinform), covering ligand-based pharmacophore (from active set alignment) and receptor-based pharmacophore (from binding pocket geometry). Explicit handling of feature types, geometric tolerances, partial matching, and pharmacophore-based virtual screening. Use when identifying scaffold-hopping candidates, building shape-and-feature search queries, or transferring SAR across chemotypes.

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Performs molecular similarity searching using Tanimoto, Tversky, Dice, and cosine coefficients on bit/count fingerprints with explicit choice rules for symmetric vs asymmetric measures, scaffold-hopping vs lead-optimization regimes, activity-cliff diagnosis, and large-library nearest-neighbor methods (BulkTanimoto, Annoy MHFP6, USRCAT). Use when ranking compounds by structural resemblance to a query, clustering libraries, finding analogs, or diagnosing activity cliffs.

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