FridrichMethod
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- ▌ Literature Search Openalex · fridrichmethod bundleQuery the OpenAlex scholarly database for research papers, authors, institutions, topics, sources, publishers, funders, geo-locations, and keywords. Use when searching academic papers, resolving DOIs, downloading open-access PDFs, finding an author's publications, aggregating bibliometric data (citation counts, h-index, impact factor), exploring the research taxonomies, or performing DOI lookups.
- ▌ Long Read Sequencing Agent · fridrichmethodAI-powered analysis of long-read sequencing data (PacBio, ONT) for structural variant detection, isoform discovery, epigenetic modifications, and de novo assembly.
- ▌ Multimodal Medical Imaging · fridrichmethod bundleAnalyzes medical images (X-ray, MRI, CT) using multimodal LLMs to identify anomalies and generate reports.
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- ▌ Time Resolved Cryoem Agent · fridrichmethodAI-powered time-resolved cryo-EM analysis for capturing protein dynamics, drug-binding kinetics, and conformational transitions for dynamics-based drug discovery.
- ▌ Tme Immune Profiling Agent · fridrichmethodComprehensive AI-powered tumor microenvironment immune profiling integrating bulk deconvolution, single-cell analysis, and spatial transcriptomics for immunotherapy biomarker discovery.
- ▌ Ucsc Conservation And Tfbs · fridrichmethod bundleFetch Evolutionary Conservation scores (phyloP, phastCons) and Transcription Factor Binding Sites (TFBS) from the UCSC Genome Browser. Use when analyzing whether genomic variants or regions are evolutionarily conserved, functionally important, or bounded by TF regulators across major projects (ENCODE, JASPAR, ReMap).
- ▌ Bio Clip Seq Clip Alignment · fridrichmethod bundleAlign CLIP-seq reads to the genome with crosslink site awareness. Use when mapping preprocessed CLIP reads for peak calling.
- ▌ Bio Phylo Tree Manipulation · fridrichmethod bundleModify phylogenetic tree structure using Biopython Bio.Phylo. Use when rooting trees with outgroups or midpoint, pruning taxa, collapsing clades, ladderizing branches, or extracting subtrees.
- ▌ Bio Reporting Figure Export · fridrichmethod bundleExports publication-ready figures in various formats with proper resolution, sizing, and typography. Use when preparing figures for journal submission, creating vector graphics for presentations, or ensuring consistent figure styling across analyses.
- ▌ Bio Workflows Clip Pipeline · fridrichmethod bundleEnd-to-end CLIP-seq analysis from FASTQ to binding sites and motif enrichment. Use when analyzing protein-RNA interactions from CLIP-based methods.
- ▌ Bio Workflows Gwas Pipeline · fridrichmethod bundleEnd-to-end GWAS workflow from VCF to association results. Covers PLINK QC, population structure correction, and association testing for case-control or quantitative traits. Use when running genome-wide association studies.
- ▌ Cart Design Optimizer Agent · fridrichmethodAI-guided CAR-T cell design for solid tumors using antigen prioritization, safety-by-design architectures, and exhaustion-resistant engineering.
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- ▌ Clinical Note Summarization · fridrichmethod bundleStructure raw clinical notes into SOAP-format summaries with explicit contradictions, missing data, and ICD-linked assessments using the provided prompt + usage script.
- ▌ Cryoem AI Drug Design Agent · fridrichmethodAI-powered integration of cryo-EM structural data with generative AI and molecular dynamics for structure-based drug design targeting flexible proteins and membrane complexes.
- ▌ Digital Twin Clinical Agent · fridrichmethodAI-powered patient digital twin creation for clinical trial simulation, treatment outcome prediction, and personalized medicine using real-world data and multi-omics integration.
- ▌ Epigenomics Methylgpt Agent · fridrichmethodAI-powered DNA methylation analysis using MethylGPT foundation models for epigenomic profiling, differential methylation detection, and cancer epigenome characterization.
- ▌ Literature Search Europepmc · fridrichmethod bundleSearch Europe PMC for scientific literature and download open-access full texts and PDFs. Retrieve full-text XML/plain text by PMCID, get citation lists and bibliography.
- ▌ Pan Cancer Multiomics Agent · fridrichmethodAI-powered pan-cancer analysis integrating genomic, transcriptomic, proteomic, and epigenomic data for cancer subtyping, driver identification, and cross-cancer pattern discovery.
- ▌ Prompt Engineering Patterns · fridrichmethod bundleThis skill should be used when the user asks to "optimize a prompt", "improve prompt performance", "design a prompt template", "write better prompts", "debug prompt issues", "use chain-of-thought", "structured prompting", "few-shot prompting", or wants to apply advanced prompt engineering patterns for production LLM applications.
- ▌ Prs Net Deep Learning Agent · fridrichmethodGeometric deep learning-based polygenic risk score prediction using PRS-Net for modeling gene interactions, enhanced disease prediction, and cross-ancestry portability.
- ▌ Variant Interpretation Acmg · fridrichmethod bundleClassifies genetic variants according to ACMG (American College of Medical Genetics) guidelines.
- ▌ Bio Phylo Tree Visualization · fridrichmethod bundleDraw and export phylogenetic trees using Biopython Bio.Phylo with matplotlib. Use when creating publication-quality tree figures, customizing colors and labels, or exporting to image formats.
- ▌ Bio Reporting Quarto Reports · fridrichmethod bundleBuild reproducible scientific documents, presentations, and websites with Quarto supporting R, Python, Julia, and Observable JS. Use when creating reproducible reports with Quarto.
- ▌ Bio Workflows Merip Pipeline · fridrichmethod bundleEnd-to-end MeRIP-seq analysis from FASTQ to m6A peaks and differential methylation. Use when analyzing epitranscriptomic m6A modifications from immunoprecipitation data.
- ▌ Bio Workflows Smrna Pipeline · fridrichmethod bundleEnd-to-end small RNA-seq analysis from FASTQ to differential miRNA expression. Use when analyzing miRNA, piRNA, or other small RNA sequencing data.
- ▌ Coagulation Thrombosis Agent · fridrichmethodAI-powered analysis of coagulation disorders, thrombosis risk prediction, anticoagulation management, and platelet function assessment using machine learning.
- ▌ Deep Visual Proteomics Agent · fridrichmethodAI-driven integration of cellular imaging, laser microdissection, and ultra-sensitive mass spectrometry for spatially-resolved single-cell proteomics.
- ▌ Human Protein Atlas Database · fridrichmethod bundleUse when you want to retrieve semi-quantitative protein expression and spatial localisation data from the Human Protein Atlas (HPA).
- ▌ Organoid Drug Response Agent · fridrichmethodAI-powered analysis of patient-derived organoid (PDO) drug screening for personalized oncology treatment selection and biomarker discovery.
- ▌ Protein Structure Prediction · fridrichmethod bundlePredicts 3D protein structures from amino acid sequences using ESMFold or AlphaFold3 (mock).
- ▌ Tumor Clonal Evolution Agent · fridrichmethodAI-powered analysis of tumor clonal architecture, subclonal dynamics, and evolutionary trajectories from multi-region sequencing and longitudinal liquid biopsy data.
- ▌ Bio Reporting Jupyter Reports · fridrichmethod bundleCreates reproducible Jupyter notebooks for bioinformatics analysis with parameterization using papermill. Use when generating automated analysis reports, running notebook-based pipelines, or creating shareable computational notebooks.
- ▌ Bio Transcription Translation · fridrichmethod bundleTranscribe DNA to RNA and translate to protein using Biopython. Use when converting between DNA, RNA, and protein sequences, finding ORFs, or using alternative codon tables.
- ▌ Chromosomal Instability Agent · fridrichmethodAI-powered analysis of chromosomal instability (CIN) signatures for cancer prognosis, immunotherapy response prediction, and therapeutic vulnerability identification.
- ▌ Computational Pathology Agent · fridrichmethod bundleAnalyze Whole Slide Images (WSI) for digital pathology, including tissue segmentation and feature extraction.
- ▌ Cytokine Storm Analysis Agent · fridrichmethodAI-powered cytokine release syndrome (CRS) and cytokine storm analysis for prediction, monitoring, and management in immunotherapy and infectious disease.
- ▌ Data Visualization Biomedical · fridrichmethodPublication-quality visualizations for biomedical and genomics data. Use when creating volcano plots, heatmaps, UMAP plots, dot plots, survival curves, forest plots, or multi-panel figures. Includes scanpy, matplotlib, seaborn, plotly workflows with journal-ready aesthetics and proper statistical annotations.
- ▌ Liquid Biopsy Analytics Agent · fridrichmethodComprehensive analysis of liquid biopsy data (ctDNA, CTCs) for cancer detection, MRD monitoring, and response tracking.
- ▌ Mpn Progression Monitor Agent · fridrichmethodAI-powered myeloproliferative neoplasm monitoring for disease progression prediction, treatment response tracking, and transformation risk assessment in PV, ET, and myelofibrosis.
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- ▌ Tcr Repertoire Analysis Agent · fridrichmethodAI-powered T-cell receptor repertoire analysis for cancer diagnosis, immunotherapy response prediction, and therapeutic TCR selection using deep learning and multi-layer ML approaches.
- ▌ Tooluniverse Variant Analysis · fridrichmethod bundleProduction-ready VCF processing, variant annotation, mutation analysis, and structural variant (SV/CNV) interpretation for bioinformatics questions. Parses VCF files (streaming, large files), classifies mutation types (missense, nonsense, synonymous, frameshift, splice, intronic, intergenic) and structural variants (deletions, duplications, inversions, translocations), applies VAF/depth/quality/consequence filters, annotates with ClinVar/dbSNP/gnomAD/CADD via ToolUniverse, interprets SV/CNV clinical significance using ClinGen dosage sensitivity scores, computes variant statistics, and generates reports. Solves questions like "What fraction of variants with VAF < 0.3 are missense?", "How many non-reference variants remain after filtering intronic/intergenic?", "What is the pathogenicity of this deletion affecting BRCA1?", or "Which dosage-sensitive genes overlap this CNV?".
- ▌ Tumor Mutational Burden Agent · fridrichmethodCalculates and harmonizes Tumor Mutational Burden (TMB) across platforms to predict immunotherapy response.
- ▌ Bio Clip Seq Clip Peak Calling · fridrichmethod bundleCall protein-RNA binding site peaks from CLIP-seq data using CLIPper, PureCLIP, or Piranha. Use when identifying RBP binding sites from aligned CLIP reads.
- ▌ Bio Primer Design Qpcr Primers · fridrichmethod bundleDesign qPCR primers and TaqMan/molecular beacon probes using primer3-py. Configure probe Tm, primer-probe spacing, and hydrolysis probe constraints for real-time PCR assays. Use when designing qPCR primers and probes.
- ▌ Bio Workflows Atacseq Pipeline · fridrichmethod bundleEnd-to-end ATAC-seq workflow from FASTQ files to differential accessibility and TF footprinting. Covers alignment, peak calling with MACS3, QC metrics, and optional TOBIAS footprinting. Use when running end-to-end ATAC-seq analysis from FASTQ to differential accessibility.
- ▌ Bio Workflows Chipseq Pipeline · fridrichmethod bundleEnd-to-end ChIP-seq workflow from FASTQ files to annotated peaks. Covers QC, alignment, peak calling with MACS3, and peak annotation with ChIPseeker. Use when processing ChIP-seq data from alignment through peak annotation.
- ▌ Bio Workflows Riboseq Pipeline · fridrichmethod bundleEnd-to-end Ribo-seq analysis from FASTQ to translation efficiency and ORF detection. Use when analyzing ribosome profiling data to study translation.
- ▌ Bio Workflows Spatial Pipeline · fridrichmethod bundleEnd-to-end spatial transcriptomics workflow for Visium/Xenium data. Covers data loading, preprocessing, spatial analysis, domain detection, and visualization with Squidpy. Use when analyzing spatial transcriptomics data.
- ▌ Molecular Glue Discovery Agent · fridrichmethodAI-powered molecular glue discovery for targeted protein degradation, enabling neo-substrate recruitment and undruggable target degradation through E3 ligase interface modulation.
- ▌ Popeve Variant Predictor Agent · fridrichmethodAI-powered genetic variant pathogenicity prediction using PopEVE deep learning model for population-aware disease variant identification and rare disease diagnosis.
- ▌ Bio Clip Seq Clip Preprocessing · fridrichmethod bundlePreprocess CLIP-seq data including adapter trimming, UMI extraction, and PCR duplicate removal. Use when preparing raw CLIP, iCLIP, or eCLIP reads for peak calling.
- ▌ Bio Genome Assembly Assembly Qc · fridrichmethod bundleAssess genome assembly quality using QUAST for contiguity metrics and BUSCO for completeness. Essential for evaluating assembly success and comparing assemblers. Use when evaluating assembly completeness and quality.
- ▌ Bio Genome Assembly Scaffolding · fridrichmethod bundleScaffold contigs into chromosome-level assemblies using Hi-C data with YaHS, 3D-DNA, SALSA2, and validate with BUSCO and contact maps. Use when scaffolding contigs to chromosome-level assemblies.
- ▌ Bio Phylo Distance Calculations · fridrichmethod bundleCompute evolutionary distances and build phylogenetic trees using Biopython Bio.Phylo.TreeConstruction. Use when creating distance matrices from alignments, building NJ/UPGMA trees, or generating bootstrap consensus trees.
- ▌ Bio Phylo Modern Tree Inference · fridrichmethod bundleBuild maximum likelihood phylogenetic trees using IQ-TREE2 and RAxML-ng. Use when inferring publication-quality trees with model selection, ultrafast bootstrap, or partitioned analyses from sequence alignments.
- ▌ Bio Primer Design Primer Basics · fridrichmethod bundleDesign PCR primers for a target sequence using primer3-py. Specify target regions, product size, melting temperature, and other constraints. Returns ranked primer pairs with quality metrics. Use when designing standard PCR primers.
- ▌ Bio Reporting Rmarkdown Reports · fridrichmethod bundleCreate reproducible bioinformatics analysis reports with R Markdown including code, results, and visualizations in HTML, PDF, or Word format. Use when generating analysis reports with RMarkdown.
- ▌ Bio Workflows Fastq To Variants · fridrichmethod bundleEnd-to-end DNA sequencing workflow from FASTQ files to variant calls. Covers QC, alignment with BWA, BAM processing, and variant calling with bcftools or GATK HaplotypeCaller. Use when calling variants from raw sequencing reads.
- ▌ Bio Workflows Multiome Pipeline · fridrichmethod bundleEnd-to-end multiome workflow for joint scRNA-seq + scATAC-seq analysis. Covers data loading, separate modality processing, and WNN integration with Seurat/Signac. Use when analyzing joint scRNA+scATAC data.
- ▌ Bio Workflows Outbreak Pipeline · fridrichmethod bundleEnd-to-end outbreak investigation from pathogen isolates to transmission networks. Orchestrates MLST typing, AMR surveillance, phylodynamic dating, and transmission inference with TransPhylo. Use when investigating disease outbreaks or tracking pathogen transmission chains.
- ▌ Bio Workflows Scrnaseq Pipeline · fridrichmethod bundleEnd-to-end single-cell RNA-seq workflow from 10X Genomics data to annotated cell types. Covers QC, normalization, clustering, marker detection, and cell type annotation. Use when analyzing single-cell RNA-seq data.
- ▌ Chip Clonal Hematopoiesis Agent · fridrichmethodAI-powered clonal hematopoiesis of indeterminate potential (CHIP) detection, risk stratification, and cardiovascular/malignancy risk prediction using genomic and clinical data.
- ▌ Hemoglobinopathy Analysis Agent · fridrichmethodAI-powered analysis of hemoglobin disorders including sickle cell disease, thalassemias, and variant hemoglobins using HPLC, electrophoresis, and molecular data.
- ▌ Multimodal Radpath Fusion Agent · fridrichmethodAI-powered multimodal diagnostic fusion integrating radiology imaging (CT/MRI/PET), digital pathology (WSI), genomics, and clinical data for comprehensive cancer diagnosis and treatment planning.
- ▌ Pytdc Therapeutics Data Commons · fridrichmethod bundleTherapeutics Data Commons (TDC) AI-ready drug discovery datasets. Curated ADME, toxicity, DTI, DDI with scaffold/cold splits, standardized metrics, molecular oracles, and ADMET benchmarks for therapeutic ML and property prediction. For chemical database queries use chembl-database-bioactivity; for featurization use molfeat.
- ▌ Tcell Exhaustion Analysis Agent · fridrichmethodAI-powered analysis of T-cell exhaustion states, epigenetic scarring, stem-like T-cell populations, and checkpoint blockade response prediction in cancer immunotherapy.
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- ▌ Bio Clip Seq Clip Motif Analysis · fridrichmethod bundleIdentify enriched sequence motifs at CLIP-seq binding sites for RBP binding specificity. Use when characterizing the sequence preferences of an RNA-binding protein.
- ▌ Bio Read Alignment Bwa Alignment · fridrichmethod bundleAlign DNA short reads to reference genomes using bwa-mem2, the faster successor to BWA-MEM. Use when aligning DNA short reads to a reference genome.
- ▌ Bio Restriction Enzyme Selection · fridrichmethod bundleSelect restriction enzymes by criteria using Biopython Bio.Restriction. Find enzymes that cut once, don't cut, produce specific overhangs, are commercially available, or have compatible ends for cloning. Use when selecting restriction enzymes for cloning or analysis.
- ▌ Bio Workflows Biomarker Pipeline · fridrichmethod bundleEnd-to-end biomarker discovery workflow from expression data to validated biomarker panels. Covers feature selection with Boruta/LASSO, classifier training with nested CV, and SHAP interpretation. Use when building and validating diagnostic or prognostic biomarker signatures from omics data.
- ▌ Bio Workflows Cytometry Pipeline · fridrichmethod bundleEnd-to-end flow cytometry workflow from FCS files to differential analysis. Orchestrates compensation, transformation, gating/clustering, and statistical testing with CATALYST/diffcyt. Use when processing flow or mass cytometry data end-to-end.
- ▌ Radiomics Pathomics Fusion Agent · fridrichmethodAI-powered multimodal fusion of radiology (CT/MRI/PET) and pathology (H&E/IHC) imaging with clinical and genomic data for comprehensive cancer diagnostics and treatment prediction.
- ▌ Spatial Transcriptomics Analysis · fridrichmethod bundleAutomated analysis pipeline for Spatial Transcriptomics (Visium, Xenium) integrating histology and gene expression.
- ▌ Bio Genome Assembly Hifi Assembly · fridrichmethod bundleHigh-quality genome assembly from PacBio HiFi reads using hifiasm with phasing support. Use when building reference-quality diploid assemblies from HiFi data, especially with trio or Hi-C phasing for fully resolved haplotypes.
- ▌ Bio Read Alignment Star Alignment · fridrichmethod bundleAlign RNA-seq reads with STAR (Spliced Transcripts Alignment to a Reference). Supports two-pass mode for novel splice junction discovery. Use when aligning RNA-seq data requiring splice-aware alignment.
- ▌ Bio Restriction Fragment Analysis · fridrichmethod bundleAnalyze restriction digest fragments using Biopython Bio.Restriction. Predict fragment sizes, get fragment sequences, simulate gel electrophoresis patterns, and perform double digests. Use when analyzing restriction digest fragment patterns.
- ▌ Bio Small Rna Seq Mirge3 Analysis · fridrichmethod bundleFast miRNA quantification with isomiR detection and A-to-I editing analysis using miRge3. Use when quantifying known miRNAs quickly or analyzing isomiR variants and RNA editing.
- ▌ Bio Workflows Microbiome Pipeline · fridrichmethod bundleEnd-to-end 16S amplicon workflow from FASTQ reads to differential abundance. Orchestrates DADA2 ASV inference, taxonomy assignment, diversity analysis, and compositional testing with ALDEx2. Use when processing 16S/ITS amplicon data.
- ▌ Bio Workflows Neoantigen Pipeline · fridrichmethod bundleEnd-to-end neoantigen discovery from somatic variants to ranked vaccine candidates. Integrates HLA typing, MHC binding prediction, pVACtools neoantigen calling, and immunogenicity scoring. Use when identifying tumor neoantigens for personalized vaccine design or checkpoint biomarkers.
- ▌ Bio Workflows Proteomics Pipeline · fridrichmethod bundleEnd-to-end proteomics workflow from MaxQuant output to differential protein abundance. Orchestrates data import, normalization, imputation, and statistical testing with MSstats or limma. Use when processing mass spectrometry proteomics.
- ▌ Simo Multiomics Integration Agent · fridrichmethodAI-powered spatial integration of multi-omics datasets using probabilistic alignment for comprehensive tissue atlas construction and cellular state mapping.
- ▌ Bio Alignment Files Bam Statistics · fridrichmethod bundleGenerate alignment statistics using samtools flagstat, stats, depth, and coverage. Use when assessing alignment quality, calculating coverage, or generating QC reports.
- ▌ Bio Data Visualization Upset Plots · fridrichmethod bundleCreate UpSet plots to visualize set intersections as an alternative to Venn diagrams using UpSetR or upsetplot. Use when comparing overlapping gene sets, peak sets, or sample groups with more than 3 sets.
- ▌ Bio Genome Intervals Bigwig Tracks · fridrichmethod bundleCreate and read bigWig browser tracks for visualizing continuous genomic data. Convert bedGraph to bigWig, extract signal values, and generate coverage tracks using UCSC tools and pyBigWig. Use when preparing coverage tracks for genome browsers or extracting signal at specific regions.
- ▌ Bio Machine Learning Atlas Mapping · fridrichmethod bundleMaps query single-cell data to reference atlases using scArches transfer learning with scVI and scANVI models. Transfers cell type labels without retraining on combined data. Use when annotating new single-cell datasets using pre-trained reference models.
- ▌ Bio Reporting Automated Qc Reports · fridrichmethod bundleGenerates standardized quality control reports by aggregating metrics from FastQC, alignment, and other tools using MultiQC. Use when summarizing QC metrics across samples, creating shareable quality reports, or building automated QC pipelines.
- ▌ Bio Systems Biology Model Curation · fridrichmethod bundleValidate, gap-fill, and curate genome-scale metabolic models using memote for quality scores and COBRApy for manual curation. Ensure models meet SBML standards and produce biologically meaningful predictions. Use when improving draft models or preparing models for publication.
- ▌ Bio Workflows Longread Sv Pipeline · fridrichmethod bundleEnd-to-end workflow for detecting structural variants from long-read sequencing data. Covers ONT/PacBio alignment with minimap2 and SV calling with Sniffles or cuteSV. Use when detecting structural variants from long reads.
- ▌ Bio Workflows Methylation Pipeline · fridrichmethod bundleEnd-to-end bisulfite sequencing workflow from FASTQ to differentially methylated regions. Covers Bismark alignment, methylation calling, and DMR detection with methylKit. Use when analyzing bisulfite sequencing data.
- ▌ Bio Workflows Multi Omics Pipeline · fridrichmethod bundleEnd-to-end multi-omics integration workflow. Orchestrates data harmonization, MOFA/mixOmics integration, factor interpretation, and downstream analysis across transcriptomics, proteomics, metabolomics, and other modalities. Use when integrating multiple omics datasets.
- ▌ Protein Sequence Similarity Search · fridrichmethod bundleSearches for homologous protein sequences using MMseqs2 (fast, default) or BLAST (comprehensive, fallback). Trigger this whenever the user provides a protein sequence or FASTA file and asks to find homologues, sequence matches, or wants to infer protein function based on sequence similarity, but not when the user wants to infer protein function based on structural similarity.
- ▌ Alphagenome Single Variant Analysis · fridrichmethod bundleAnalyzes genetic variant effects on gene expression (RNA-seq), chromatin accessibility (DNASE), histone marks (ChIP), and transcription factors using the AlphaGenome API. Use when the user asks about non-coding variant effects, pathogenicity, clinical significance, disease associations, functional effects, gene expression changes, splicing disruption, or regulatory effects in promoters and enhancers. Also use for resolving biological terms to tissue/cell-type ontologies (UBERON/CL) or analyzing variants in chr:pos:ref>alt format.
- ▌ Bio Data Visualization Circos Plots · fridrichmethod bundleCreate circular genome visualizations with Circos and pyCircos. Display multi-track data including ideograms, genes, variants, CNVs, and interaction arcs. Use when creating circular genome visualizations.
- ▌ Bio Experimental Design Sample Size · fridrichmethod bundleEstimates required sample sizes for differential expression, ChIP-seq, methylation, and proteomics studies. Use when budgeting experiments, writing grant proposals, or determining minimum replicates needed to achieve statistical significance for expected effect sizes.
- ▌ Bio Expression Matrix Counts Ingest · fridrichmethod bundleLoad gene expression count matrices from various formats including CSV, TSV, featureCounts, Salmon, kallisto, and 10X. Use when importing quantification results for downstream analysis.
- ▌ Bio Primer Design Primer Validation · fridrichmethod bundleValidate PCR primers for specificity, dimers, hairpins, and secondary structures using primer3-py thermodynamic calculations. Check self-complementarity, heterodimer formation, and 3' stability. Use when validating primer specificity and properties.