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Detects differential alternative splicing between conditions using rMATS-turbo (binomial LRT on junction counts), leafcutter (Dirichlet-multinomial GLM on intron clusters), MAJIQ V3 deltapsi/HET (Bayesian posterior on LSVs), SUPPA2 (empirical-null on TPM-derived PSI), or Shiba (junction-imbalance-corrected, 2025 SOTA at low coverage). Reports FDR-corrected significance and delta PSI effect sizes. Tools differ in statistical model, annotation dependence, calibration regime, and replicate-count requirements. Use when comparing splicing patterns between treatment groups, tissues, or disease states.

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Perform multiple sequence alignment using MAFFT, MUSCLE5, ClustalOmega, or T-Coffee. Guides tool and algorithm selection based on dataset size, sequence divergence, and downstream application. Use when aligning three or more homologous sequences for phylogenetics, conservation analysis, or evolutionary studies.

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Quantifies alternative splicing as PSI (percent spliced in) from RNA-seq using rMATS-turbo (BAM-based event), SUPPA2 (TPM-based event), MAJIQ V3 (LSV-based Bayesian), leafcutter (annotation-free intron clusters), VAST-TOOLS (cross-species with microexon support), Shiba (junction-imbalance-corrected, 2025 SOTA at low coverage), or IRFinder-S (intron retention coverage-aware). Distinguishes the five canonical event classes (SE, A5SS, A3SS, MXE, RI), special classes (microexons, exitrons, AFE/ALE), intron retention subtypes (canonical RI vs detained introns), and applies effective-length normalization. Use when measuring splice-site usage or isoform inclusion ratios from short-read RNA-seq.

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View, convert, and understand SAM/BAM/CRAM alignment files using samtools and pysam. Use when inspecting alignments, converting between formats, or understanding alignment file structure.

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Generate consensus sequences and manage reference files using samtools. Use when creating consensus from alignments, indexing references, or creating sequence dictionaries.

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Generate pileup data for variant calling using samtools mpileup and pysam. Use when preparing data for variant calling, analyzing per-position read data, or calculating allele frequencies.

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Sort alignment files by coordinate or read name using samtools and pysam. Use when preparing BAM files for indexing, variant calling, or paired-end analysis.

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Mark and remove PCR/optical duplicates using samtools fixmate and markdup. Use when preparing alignments for variant calling or when duplicate reads would bias analysis.

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Trim PCR primers from aligned reads in amplicon-panel BAMs using samtools ampliconclip. Use when processing SARS-CoV-2 ARTIC, hereditary cancer panels, ctDNA hot-spot panels, or any amplicon assay where primer-derived bases would falsely confirm reference at primer footprints.

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Analyzes alternative splicing at single-cell resolution. The first decision is library chemistry — 10X 3' is fundamentally limited (RT primes from poly-A, R2 falls in 3' UTR, <0.1 junction read per cell per AS event). Plate-based full-length methods (Smart-seq3, FLASH-seq, VASA-seq, STORM-seq) and single-cell long-read (MAS-Iso-seq, scISOr-Seq2) are the chemistries that give per-cell isoform structure. Tools include MARVEL (R, Smart-seq integrated), BRIE2 (Bayesian PSI with regulatory features and ELBO_gain test), scQuint (junction-cluster, plate-based; not for 10X), SpliZ (annotation-free Z-score), Psix (graph-smoothness regulated AS), and Sierra (alternative polyadenylation, often confused with AS). Use when analyzing isoform usage in scRNA-seq, identifying cell-type-specific splicing, or determining whether scRNA-seq chemistry supports splicing analysis at all.

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Create and use BAI/CSI indices for BAM/CRAM files using samtools and pysam. Use when enabling random access to alignment files or fetching specific genomic regions.

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Generate alignment statistics using samtools flagstat, stats, depth, coverage, and mosdepth. Use when assessing alignment quality, calculating coverage, or generating QC reports.

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Build low-fidelity wireframes, screen mockups, and user-flow diagrams in Frame0 (the hand-drawn Balsamiq-style wireframing tool) through its connected MCP server. Use when asked to wireframe a screen, sketch a UI, make a low-fi mockup, lay out an app's screens, or diagram a user flow with clickable links. Triggers: "wireframe", "frame0", "low-fi mockup", "sketch a screen", "user flow", "lo-fi", "rough out the UI". NOT for high-fidelity visual design, brand systems, or color/typography polish (use web-design-expert / Figma); NOT for production UI code (use frontend skills).

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Master of text-based visual communication using ASCII art, Unicode box-drawing, and structured diagram notation. Creates clear, maintainable diagrams for systems, processes, hierarchies, relationships, and psychological structures. Proactively generates diagrams to enhance understanding. Activate on visualization needs, system architecture, process flows, psychological mapping, or when complex concepts would benefit from visual representation. NOT for photo editing, vector graphics, or GUI-based design tools.

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Choose, compare, and decompose work into the right coordination topology: dag, workflow, manager-driven team, swarm, blackboard, team-builder, or recurring loop. Use when the coordination shape is unclear, when you need topology-native breakdown instructions instead of a generic DAG, or when planning topology and runtime topology may differ. NOT for actual execution, node-level skill matching, or framework/tool installation.

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Generates shell completion scripts for CLI tools. Parses command structure from help output, commander/yargs configs, or manual specification. Produces completions for bash, zsh, and fish that follow each shell's conventions.

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Expert in cluttering -- the lesser-known fluency disorder characterized by rapid/irregular speech rate, excessive disfluencies, and frequent ADHD comorbidity. Covers differential diagnosis from stuttering, assessment protocols, therapy approaches (rate control, self-monitoring, pragmatic awareness), technology-assisted intervention, and AI tools for people who clutter. Activate on 'cluttering', 'rapid speech disorder', 'speech rate too fast', 'speech disfluency not stuttering', 'disorganized speech', 'ADHD speech problems', 'cluttering vs stuttering', 'speech intelligibility', 'mazed speech', 'telescoping words'. NOT for stuttering-only treatment (overlap is common but they are distinct), hearing disorders, or voice disorders (pitch, resonance).

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Audit whether an enforced-coordination MCP has actually shrunk its ~19 legacy etiquette verbs into the 5 enforced tools — work/act/ask/recall/status — each carrying one denial shape and one transcript event, migrated through a real retirement path (intake-metadata, alias, or doc-history — never a parallel runtime), with zero legacy-verb calls once a body reaches compliance mode C4+. Use when reviewing a broker-collapse migration plan, gating a body's move from advisory (C0-C3) to enforced (C4+) coordination, or verifying IT-018 Broker Collapse compliance. NOT for the inbound trigger→spawn boundary that decides whether an event may spawn an agent at all (use fleet-event-spawn-trust), the capability-token cryptographic mechanism behind a denial (use agentic-zero-trust-security), designing how a swarm of agents invokes and shards work (use swarm-invocation-designer), or auditing an individual PR's mergeability (use agent-pr-authoring).

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Cooperative vibe coding across machines — real-time collaborative development with humans and AI agents. Covers current tools (Live Share, Code With Me, Zed, Cursor multiplayer, tmux), git-based async patterns, AI agent coordination, and the aspirational future of multi-human multi-agent development. Activate on: "pair programming", "vibe coding together", "collaborative coding", "mob programming", "remote pairing", "code together", "multiplayer coding", "Live Share", "Code With Me", "ensemble programming", "cooperative development". NOT for: building a real-time collaboration ENGINE (use real-time-collaboration-engine), git workflow mechanics (use git-best-practices), single-user AI coding assistance.

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Send and receive transactional emails with Cloudflare Email Service (Email Sending + Email Routing). Use when building email sending (Workers binding or REST API), email routing, Agents SDK email handling, or integrating email into any app — Workers, Node.js, Python, Go, etc. Also use for email deliverability, SPF/DKIM/DMARC, wrangler email setup, MCP email tools, or when a coding agent needs to send emails. Even for simple requests like "add email to my Worker" — this skill has critical config details.

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Audits an Articles of Agreement contract — the daemon-witnessed agreement every official Port Daddy agent signs covering registration, transcript reporting, tool-use gating, file claims, parley conduct, budget limits, and operator control — against the enforcement-beats-hope bar: a clause counts as safe only when it resolves to a concrete, daemon-observable mechanism with a defined denial shape, never a promise the agent might honor. Use when drafting a new agent's Articles, reviewing a compliance-level (C0-C6) claim, deciding whether a clause is genuinely enforced or merely documented, or auditing whether an agent's claimed identity can be trusted. NOT for securing the inbound event-to-spawn path (fleet-event-spawn-trust), designing the relay's transport PKI and signing keys (pd-relay-zero-trust), or tracking an agent's identity and reputation across its whole lifetime after it signs (agent-identity-continuity-reputation).

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Design decisions for when and what sounds to use in web apps, mobile apps, and desktop native apps — plus how to create them with AI. Covers UI sound vocabulary (confirms, alerts, transitions, errors), platform conventions (iOS, macOS, Material), creating sounds with AI tools (ElevenLabs, AudioCraft, Stable Audio), Web Audio API for procedural sounds, haptic-audio pairing, and the psychology of satisfying vs. annoying UI sounds. Activate on 'app sounds', 'UI sounds', 'notification sound', 'sound design app', 'earcons', 'sonic branding', 'interaction sounds', 'feedback sounds', 'app audio', 'satisfying sounds'. NOT for professional audio engineering (use sound-engineer), Windows 3.1 retro sounds specifically (use win31-audio-design), music composition (use sound-engineer).

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Refactor apps so backend owns runtime truth and UI only projects snapshots. Use when tools have split authority. NOT for cosmetic UI work or new features before drift is removed.

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Design RL simulation sandboxes, trajectory datasets, QLoRA/LoRA adaptation plans, eval harnesses, and rollback unhooks for agents learning specific coding behaviors beyond the base model. Use when building or reviewing tool-use training loops, offline trajectories, reward functions, self-improvement harnesses, or Port Daddy agent behavior curricula. NOT for generic ML tutorials, full model training operations, or production deployment without eval gates and safety unhooks.

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