查找技能

Reads, queries, and writes bigWig indexed binary signal tracks (coverage, fold-change, conservation, methylation-rate) with pyBigWig (Python) and the UCSC Kent tools (bedGraphToBigWig, bigWigToBedGraph, bigWigInfo, bigWigSummary, bigWigAverageOverBed) and deepTools (multiBigwigSummary, computeMatrix, bigwigCompare). Covers the central trap that a wide query returns a precomputed zoom-level summary (by default the mean, which annihilates narrow peaks) not per-base data, when exact=True/values() is mandatory, the NaN-not-zero gap-handling fork, choosing mean vs max vs sum vs coverage by biological question, and the sorted-bedGraph plus chrom.sizes build requirement. Use when extracting signal at regions, computing mean signal per gene/peak, building a browser track from bedGraph, comparing tracks, or building TSS/gene-body metaprofiles.

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Segment single cells from multiplexed IMC/MIBI tissue images using Mesmer/DeepCell, Cellpose, or ilastik+CellProfiler, covering whole-cell vs nuclear segmentation, the summed-membrane-channel decision, nuclear-expansion bias, lateral spillover, resolution-floor parameters, and downstream-proxy evaluation. Use when delineating cells after preprocessing, choosing a segmentation model, building a cell mask for quantification, diagnosing impossible double-positive populations, or troubleshooting over/under-segmentation.

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Detects and removes contamination in genome assemblies via two disjoint workflows - foreign-sequence screening of a single-organism (eukaryote/isolate) assembly with NCBI FCS-GX (GenBank-submission-mandatory), FCS-adaptor, and BlobToolKit blob plots; and MAG/bin quality assessment with CheckM2 plus GUNC (chimerism) plus GTDB-Tk taxonomy, judged against MIMAG. Covers why CheckM2 alone is blind to disjoint-marker chimeras, the FCS-GX RAM wall, organelle/NUMT triage, strain heterogeneity, and the HGT-vs-contamination (tardigrade) trap. Use when screening an assembly for foreign contamination before GenBank submission, assessing MAG completeness/contamination/chimerism, deciding which contigs to remove, or distinguishing real HGT from contaminant contigs.

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Generates, normalizes, and converts bedGraph signal tracks (4-column chrom/start/end/value, 0-based half-open) with bedtools genomecov, deepTools bamCoverage/bamCompare/bigwigCompare, bedtools unionbedg, and UCSC bedGraphToBigWig. Covers why a raw coverage bedGraph is not comparable across samples until normalized, the CPM/RPKM/BPM/RPGC normalization menu and the conserved-total assumption that makes them wrong under a global perturbation, the strict sorted-non-overlapping-chrom.sizes bedGraphToBigWig contract that silently corrupts a bigWig, effective-genome-size selection, and bin-size aliasing. Use when building or normalizing a coverage/signal track from a BAM, comparing tracks across samples or conditions, converting bedGraph to a browser-ready bigWig, or diagnosing a track that looks plausible but reports wrong heights.

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Detects A/B chromatin compartments from balanced Hi-C contact matrices via eigenvector decomposition of the distance-normalized, Pearson-correlated cis matrix with cooltools (eigs_cis), then orients (phases) the compartment eigenvector against a GC or gene-density track so the active (A) sign is not arbitrary. Covers the eigenvector-is-a-choice problem (per-arm view_df to remove the centromere gradient; picking the eigenvector by max correlation with activity, not by eigenvalue), GC phasing with bioframe.frac_gc, resolution choice (100kb-1Mb), saddle plots and saddle_strength for compartmentalization strength, the cohesin-loss-strengthens-compartments result, subcompartments (SNIPER/Calder/dcHiC), and cross-condition compartment switching. Use when calling A/B compartments, computing E1/eigenvectors, phasing the eigenvector, building saddle plots, choosing a compartment resolution, quantifying compartment strength, or comparing compartmentalization across conditions.

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Calls significant loops from protein-directed and targeted 3C assays (HiChIP, PLAC-seq, Capture Hi-C/PCHi-C, ChIA-PET) where the contact background is peak-anchored and coverage-biased, so generic Hi-C loop callers (cooltools dots, Juicer HiCCUPS) use the wrong null. Covers FitHiChIP (config-driven coverage+distance-decay spline regression, peak-to-peak vs peak-to-all foreground, loose vs stringent background, coverage vs ICE bias), MAPS (positive Poisson regression on bias factors for PLAC-seq/HiChIP), hichipper (restriction-site-distance bias model + library QC), CHiCAGO (Delaporte two-component Brownian+technical background for asymmetric bait x other-end Capture Hi-C), the with/without separate-ChIP anchor decision, and differential loops via diffloop. Use when calling loops from HiChIP/PLAC-seq/Capture Hi-C, choosing FitHiChIP/MAPS/CHiCAGO, picking peak-to-all vs peak-to-peak, setting the loop FDR, supplying ChIP peaks as anchors, QCing a HiChIP library, or comparing loops between conditions.

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Assigns GO terms, Pfam/InterPro domains, KEGG orthologs, EC numbers, and product names to predicted proteins using eggNOG-mapper (orthology), InterProScan (domain signatures), and KofamScan (KEGG), routing specialized functions to dbCAN/antiSMASH/AMRFinderPlus/SignalP. Covers the orthology-vs-domain-vs-homology paradigms, the annotation-error percolation cascade, domain-presence-is-not-function, GO IEA circularity in enrichment, evidence tiering, and bit-score/coverage thresholds. Use when adding functional annotation to predicted genes, choosing between eggNOG-mapper and InterProScan, or judging how much to trust a functional label.

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Decides whether and how to polish a draft genome assembly to raise consensus accuracy (QV) with read-type-matched tools - Racon and medaka (ONT consensus), dorado polish, Polypolish and pypolca (Illumina, repeat-aware), Pilon (legacy short-read), NextPolish/NextPolish2, Hapo-G (haplotype-aware), ntEdit, and DeepPolisher/PEPPER-Margin-DeepVariant for human. Covers the do-not-polish-HiFi rule, the medaka basecaller-model footgun, held-out Merqury QV as the only honest stop signal, and the haplotype-collapse trap. Use when correcting homopolymer indels or residual SNPs in a long-read assembly, deciding if a HiFi assembly needs polishing, or choosing an ONT vs hybrid vs short-read polishing chain.

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Orders and orients assembled contigs into chromosome-scale scaffolds from long-range linking data, inserting N-gap spacers (adds no sequence). Covers Hi-C/Omni-C scaffolding (YaHS, SALSA2, 3D-DNA/Juicer), Hi-C read-mapping prerequisites (map each end separately, no mate rescue, dedup, enzyme-aware), reading the contact map for misjoins/inversions/false-duplications, manual curation in Juicebox/PretextView (the VGP/DToL standard), reference-guided scaffolding (RagTag) and its karyotype-erasure hazard, genetic-map (ALLMAPS) and Bionano optical-map integration, chimera-breaking before scaffolding, gap-filling, and telomere/contig-vs-scaffold-N50 QC (tidk). Use when turning contigs into chromosomes with Hi-C, integrating a linkage map or optical map, choosing a scaffolder by available linking data, or judging whether a chromosome-scale assembly is trustworthy.

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Handles BED-format genomic intervals (BED3 through BED12, narrowPeak/broadPeak) and the coordinate-system substrate the whole interval category rests on, with bedtools (CLI) and pybedtools/pyranges/pandas (Python). Covers the 0-based half-open vs 1-based-closed convention boundary and the start-1/end-unchanged conversion, the silent failures (chrom-name mismatch, CRLF, lexicographic-vs-version sort under -sorted), genome/chrom.sizes generation, sorting contracts, BED12 block invariants, validation, makewindows, cross-assembly liftover (liftOver/CrossMap), and BED<->VCF/BAM/FASTA conversion. Use when reading, creating, validating, sorting, lifting between genome builds, or converting interval files, preparing inputs for bedtools/tabix/bigBed, or debugging an off-by-one or empty-overlap result.

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Tests whether two genomic interval sets overlap (colocalize) more than expected by chance using a permutation test against a structured-genome null model. Covers bedtools fisher (analytic 2x2 screen), bedtools shuffle + jaccard permutation, GAT (isochore/GC-conditioned simulation with FDR), regioneR (flexible permutation, randomizeRegions vs circularRandomizeRegions, localZScore), LOLA (universe-relative Fisher against a region database), and GREAT/rGREAT (regulatory-domain binomial + hypergeometric for ontology-from-regions). Stresses the universe/background choice, matched background, blacklist exclusion, and multiple-testing control. Use when asking whether peaks/regions are enriched at enhancers/TFBS/features, scoring region-set colocalization or region-set enrichment, comparing CNV/SV concordance, or turning an overlap count into a defensible p-value.

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Performs proximity operations on genomic intervals with bedtools (closest, window, flank, slop) and pybedtools - nearest-feature queries with signed/strand-aware distance, fixed-radius window searches, strand-aware promoter construction, and interval extension. Covers the closest -d/-D a/b/ref/-t/-k/-io/-iu/-id flags, the -D ref strand sign-flip, silent chromosome-end clipping in slop/flank, -t all tie double-counting, and the critical distinction between a geometry answer (nearest TSS) and a biology answer (which gene an element regulates). Use when assigning peaks or variants to genes, defining promoters from a gene model, building distance-to-TSS distributions, finding features within a window, or extending intervals - and when deciding whether nearest-gene is a fair prior (GWAS locus) or a trap (distal enhancer).

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Turns Hi-C/Micro-C FASTQ into a deduplicated, filtered .pairs file with pairtools and decides whether the library worked. Covers the bwa mem -SP5M / bwa-mem2 / chromap --preset hic alignment idiom (mates mapped as independent single-end reads), pairtools parse vs parse2 and the walks-policy choice (5unique pairwise vs all for Pore-C/Micro-C concatemers), pair-type classification (keep UU and rescued UC), dedup (PCR vs optical/by-tile), select by pair_type/MAPQ/distance, restriction-fragment handling (restrict, Arima dual-enzyme, Micro-C/DNase fragment-free), and allele-specific phasing (pairtools phase to two coolers). The library-QC decision uses % long-range cis as the one-number quality metric, trans as the noise floor, orientation balance as fragment-map-free dangling-end/self-circle QC, and % duplicates as a complexity proxy. Use when processing Hi-C/Micro-C/Omni-C reads into pairs, judging library quality, handling multi-enzyme or restriction-agnostic protocols, or generating allele-specific contacts.

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Renders Hi-C contact matrices honestly and reproducibly with matplotlib, cooltools, HiCExplorer, pyGenomeTracks, FAN-C, CoolBox, and plotgardener. Covers the raw/ICE-balanced/observed-over-expected transform choice, LogNorm vs symmetric-diverging colormaps with vmax/percentile clipping, resolution-to-feature matching (compartments 100-500kb, TADs 10-40kb, loops 5-10kb), square vs rotated-triangle track-stacking, NaN/white-stripe handling, virtual 4C, APA/saddle/on-diagonal pileups, two-condition side-by-side and log2-ratio maps, and interactive (HiGlass) vs scripted-static publication figures. Use when plotting a contact matrix, choosing a normalization or color scale, building a multi-track Hi-C figure, making a virtual 4C profile, piling up loops/boundaries, or comparing two conditions.

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Analyze spatial cell-cell interactions, neighborhoods, and niches in IMC/MIBI data with squidpy and imcRtools, covering neighborhood-enrichment permutation nulls, the abundance-vs-density confound, inhomogeneous Ripley's K, cellular-neighborhood discovery, graph-construction (contact vs proximity), and edge effects. Use when testing whether cell types co-locate, choosing a spatial null, building a neighbor graph, discovering tissue niches, or deciding whether a spatial pattern is real or a density/segmentation artifact.

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Designs cytosine (CBE, C-to-T) and adenine (ABE, A-to-G) base-editor guides by positioning the target base at the activity-peak of the editing window (protospacer positions ~5-7, PAM-distal numbering), minimizing bystander edits for product purity, reading dinucleotide context (APOBEC1 TC favored / GC disfavored), and selecting the editor variant (BE4max, ABEmax, ABE8e, YE1/SECURE, TadCBE, CGBE, SpG/SpRY-BE). Covers knockout by premature stop (CRISPR-STOP/iSTOP) and splice-site disruption, the three off-target classes (Cas-dependent, Cas-independent DNA, RNA), outcome prediction (BE-Hive/DeepBE), and the base-vs-prime-vs-HDR decision. Use when installing a transition mutation without a double-strand break, knocking out a gene without indels, or choosing CBE vs ABE. Generic guide scoring, prime editing, and HDR donors are separate skills.

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Predict peptide-MHC class I binding and natural presentation with MHCflurry, NetMHCpan-4.1, and MixMHCpred to nominate candidate CD8 T-cell epitopes. Covers the binding-affinity (BA) vs eluted-ligand (EL/presentation) distinction, why %Rank beats raw nM for cross-allele work, the MS abundance bias that misranks low-expression neoantigens, allele-coverage inequity, and length bias. Use when scanning a protein or peptide set for class I epitopes, scoring neoantigen candidates, or choosing a binding predictor. For CD4/HLA class II see mhc-class-ii-prediction.

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Discovers, classifies, and masks repetitive elements and transposable elements with RepeatModeler2 (de novo family library), RepeatMasker (masking against a library), EDTA (plant/structural TEs), or EarlGrey (auto-curating wrapper), and quantifies TE expression from RNA-seq with TEtranscripts/SQuIRE. Covers de-novo-library-as-curation-project, soft-vs-hard masking, the domesticated-gene over-masking massacre, Dfam-vs-RepBase, TE classification (Class I/II, family-vs-copy), Kimura repeat landscapes, LAI, and the RNA-seq multimapping problem. Use when masking repeats before gene prediction, building a TE library for a non-model genome, or analyzing transposable-element content or expression.

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Assembles haplotype-resolved diploid and telomere-to-telomere (T2T) genomes from PacBio HiFi reads with hifiasm (HiFi-only, Hi-C, or trio phasing) and verkko (HiFi + ultralong ONT for T2T), extracting contigs from GFA and routing phasing QC to k-mer/trio metrics. Covers why a primary assembly is a haplotype mosaic that exists in no cell, partial-vs-full phasing (the .bp. vs .dip. filename convention), the purge-default trap on inbred samples, the --hom-cov coverage-estimate alarm, and verkko-vs-hifiasm for T2T. Use when assembling a diploid eukaryote from HiFi, phasing haplotypes with parents (trio) or Hi-C, deciding whether to chase T2T, or diagnosing switch errors invisible to N50/BUSCO/QV.

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Assembles genomes de novo from noisy long reads (Oxford Nanopore R9/R10/Dorado, PacBio CLR) with Flye (repeat graph), Canu (correct-trim-assemble OLC), NextDenovo, Shasta, Raven, wtdbg2, or miniasm, and reconciles bacterial assemblies into a consensus with Trycycler/Autocycler. Covers matching the input flag to the basecaller era (--nano-hq vs --nano-raw), why a raw long-read assembly is contiguous but low-QV and not finished until polished, haplotig false-duplication and purge_dups, coverage and read-N50 as non-substitutable inputs, and mid-read adapter de-chimerization. Use when assembling a bacterial or eukaryotic genome from ONT or PacBio noisy reads, choosing a long-read assembler, or diagnosing an over-collapsed or duplicated assembly. For PacBio HiFi use hifi-assembly instead.

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Assembles a genome de novo from Illumina short reads with SPAdes (isolate/careful/sc/meta/plasmid/rna modes), MEGAHIT (low-memory, huge datasets), Unicycler (bacterial finishing/hybrid), MaSuRCA (large hybrid), ABySS (Bloom-filter), and Platanus (heterozygous diploids), using multi-k de Bruijn graphs. Covers the repeat-resolution limit, why N50 plateaus at the genome not the depth, GenomeScope2 k-mer profiling first, the heterozygosity/haplotig trap, error-correction erasing rare alleles, GC dropout, and NG50/auN/BUSCO reporting. Use when assembling a bacterial isolate, fungal, small-eukaryotic, single-cell, or metagenome genome from Illumina reads, or when deciding whether short reads can even produce the assembly being asked for.

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Designs and ranks guide RNAs (sgRNAs) for CRISPR-Cas9/Cas12a gene knockout by scanning a target for PAM sites (NGG SpCas9, NNGRRT SaCas9, TTTV Cas12a, NG SpCas9-NG, near-PAMless SpRY), enumerating candidate spacers, applying hard filters (Pol-III TTTT terminator, 5' G, GC), ranking on-target activity with the context-appropriate model (Rule Set 2/Azimuth for U6/lentiviral, CRISPRscan for T7/embryo, DeepHF for high-fidelity variants, DeepCpf1 for Cas12a), and predicting the indel/frameshift outcome (Bae out-of-frame score, inDelphi, FORECasT, Lindel). Use when selecting sgRNAs to knock out a gene, choosing a nuclease/PAM for a constrained locus, picking which exon to target, or shortlisting guides before an off-target check. Off-target specificity, base/prime editing, and HDR donors are separate skills.

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Computes and interprets sequencing read depth and coverage over a genome, windows, or target regions with mosdepth (windowed depth, cumulative distribution, --quantize callable BEDs), bedtools genomecov/coverage (bedGraph tracks, per-target stats), samtools depth/coverage (per-base depth, per-contig depth+breadth). Covers the breadth-vs-mean distinction, the cumulative-coverage curve, evenness (CV/Fano/fold-80/Gini), what each tool silently counts (duplicates, secondary/supplementary, MAPQ, read span vs fragment, mate-overlap), the samtools-depth 8000-cap version trap, and the bedtools coverage -a/-b orientation flip. Use when assessing sequencing adequacy, building coverage tracks, computing breadth at a depth threshold, defining callable regions, or QCing target-capture uniformity.

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Compare cell-type composition and spatial features across conditions in IMC/MIBI cohorts with the patient as the experimental unit, covering pseudoreplication, per-patient aggregation, mixed models, compositional (Dirichlet/scCODA) differential abundance, diffcyt, per-image-to-patient spatial differential testing (SpaceANOVA), batch covariates, and FDR. Use when testing whether a cell type or spatial niche differs between groups, avoiding cell-level pseudoreplication, choosing a differential-abundance method, or correctly powering an IMC cohort comparison.

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