Medical Genetics

DNA: Double helix (B-form). Nucleotides: phosphate + deoxyribose + nitrogenous base (purines: adenine, guanine; pyrimidines: cytosine, thymine). Base pairing: A=T (2 H-bonds), G≡C (3 H-bonds). Antiparallel strands (5'→3' and 3'→5'). Major and minor grooves for protein binding. Chromatin: DNA wrapped around histones (H2A, H2B, H3, H4) forming nucleosomes, 30nm fiber, loops, and chromosomes. RNA: Single-stranded, ribose instead of deoxyribose, uracil instead of thymine. Types: mRNA (messenger, protein-coding), rRNA (ribosomal, structural/catalytic), tRNA (transfer, amino acid delivery), snRNA (splicing), miRNA/siRNA (gene silencing), lncRNA (regulatory). Telomeres: TTAGGG repeats at chromosome ends. Telomerase (reverse transcriptase + RNA template) maintains telomeres in stem cells/germ cells. Shortening linked to aging and cellular senescence.

Semiconservative replication (Meselson-Stahl). Origins of replication: Multiple origins (eukaryotes), ORC complex. Replication forks bidirectionally. Enzymes: DNA helicase (MCM complex) unwinds DNA, SSB proteins stabilize single strands, topoisomerase (Topo I/II) relieves supercoiling. Primase synthesizes RNA primers. DNA polymerase α (primer extension), δ (leading), ε (lagging) in eukaryotes. DNA polymerase III (prokaryotes). 5'→3' polymerase activity, 3'→5' exonuclease (proofreading). Leading strand (continuous), lagging strand (Okazaki fragments). RNase H removes RNA primers, DNA ligase seals nicks. Telomerase: Adds TTAGGG repeats to 3' end of lagging strand. Active in germ cells, stem cells, cancer cells. Cell cycle: G1 (growth), S (DNA synthesis), G2 (growth/preparation), M (mitosis: prophase, metaphase, anaphase, telophase). Checkpoints: G1/S (Rb/p53), G2/M (cyclin B/CDK1), M (spindle checkpoint).

RNA Polymerases: RNA Pol I (rRNA), RNA Pol II (mRNA, snRNA, miRNA, lncRNA), RNA Pol III (tRNA, 5S rRNA, U6 snRNA). Promoters: Core promoter (TATA box at -30, initiator element, downstream promoter element). Proximal promoter elements (CAAT box, GC box). Enhancers (distal, tissue-specific, orientation-independent). Silencers, insulators/boundary elements. Transcription factors: Basal (TFIIA, TFIIB, TFIID/TBP, TFIIE, TFIIF, TFIIH). Mediator complex bridges activators to Pol II. Steps: Initiation (closed complex, open complex, promoter clearance), elongation (Pol II CTD phosphorylated by TFIIH/CDK7, elongation factors P-TEFb, DSIF, NELF), termination (polyadenylation signal AAUAAA, CPSF/CstF cleavage, Rat1/Xrn2 torpedo model). Pre-mRNA processing: 5' capping (7-methylguanosine, protects from exonucleases, enhances translation), splicing (spliceosome: U1, U2, U4/U6, U5 snRNPs, branch point A, 5' splice site GU, 3' splice site AG, lariat intermediate, alternative splicing), 3' polyadenylation (poly-A polymerase adds ~200 As, enhances stability/export). RNA editing: Deamination of A to I (ADAR), C to U (APOBEC).

Ribosome: Eukaryotic 80S (40S small + 60S large). Prokaryotic 70S (30S + 50S). tRNA: Cloverleaf structure, anticodon loop, acceptor stem (3' CCA + amino acid). Aminoacyl-tRNA synthetases charge tRNA (one per amino acid, ATP-dependent). Wobble hypothesis: 3rd base flexibility allows 61 codons to be read by ~45 tRNAs. Genetic code: 61 sense codons (20 amino acids + start AUG/methionine), 3 stop codons (UAA, UAG, UGA). Degenerate, universal (exceptions: mitochondria). Initiation: eIF4E binds 5' cap, eIF4G/eIF4A, scanning to AUG (Kozak sequence ACCAUGG). eIF2-GTP-Met-tRNAi ternary complex. 60S joins, eIFs released. Elongation: eEF1A delivers aminoacyl-tRNA, peptidyl transferase (28S rRNA, ribozyme) forms peptide bond. eEF2 translocates ribosome. Termination: eRF1 recognizes stop codons, eRF3 GTPase, polypeptide released. Ribosome recycling. Post-translational: Folding (chaperones Hsp70, Hsp90), glycosylation (ER/Golgi), phosphorylation, ubiquitination (proteasome), signal sequences (SRP, ER targeting), cleavage (proteases).

Transcriptional regulation: Promoters, enhancers (looping via Mediator/cohesin), silencers, insulators (CTCF). Chromatin states: euchromatin (active, open) vs heterochromatin (inactive, condensed, constitutive H3K9me3/H4K20me3, facultative). Histone modifications: Acetylation (HATs: active; HDACs: repressive). Methylation (H3K4me3 = active promoters; H3K9me3 = heterochromatin; H3K27me3 = Polycomb repression; H3K36me3 = transcriptional elongation). Phosphorylation, ubiquitination, SUMOylation. Histone code hypothesis. DNA methylation: CpG islands (promoters, mostly unmethylated in active genes). DNMT3A/3B (de novo), DNMT1 (maintenance). Methylation = transcriptional silencing. Imprinted genes, X-inactivation, retrotransposon silencing. Chromatin remodeling: SWI/SNF (BAF complex, mobilizes nucleosomes, ATP-dependent), ISWI, CHD, INO80. Mutations in BAF subunits (SMARCB1, ARID1A, SMARCA4) in cancers. Non-coding RNAs: miRNA (22nt, Dicer processing, RISC complex, binds 3'UTR, translational repression/mRNA decay). siRNA (exogenous/viral, perfectly complementary, RNAi pathway). lncRNA (XIST, HOTAIR, MALAT1, NEAT1, dosage compensation, scaffold, guide, decoy). piRNA (germline, transposon silencing). CRISPR-Cas9: Adaptive bacterial immune system. Cas9 endonuclease + guide RNA (crRNA + tracrRNA). DSB at target site: NHEJ (indels, gene disruption) or HDR (with donor template, precise editing). Applications: gene knockout, knock-in, activation (CRISPRa), repression (CRISPRi), base editing (deaminase), prime editing (reverse transcriptase), epigenetic editing. Clinical trials for sickle cell (exa-cel/Casgevy), T-cell engineering (CAR-T), DMD, LCA10.

Caused by meiotic nondisjunction (increases with maternal age). Most aneuploidies are lethal except for specific trisomies and sex chromosome abnormalities. Trisomy 21 (Down syndrome): 1:700 live births. Features: intellectual disability, brachycephaly, flat facies, epicanthal folds, Brushfield spots, single palmar crease, sandal gap, duodenal atresia, AVSD/VSD, Hirschsprung disease, increased AML/ALL risk (20x), early Alzheimer disease (APP gene on chr21). 95% full trisomy (meiosis I nondisjunction), 4% Robertsonian translocation t(14;21), 1% mosaic. Recurrence risk: 1% for full trisomy, 10-15% for maternal t(14;21) carrier, <1% for paternal carrier. Trisomy 18 (Edwards): 1:5000. Rocker-bottom feet, clenched hands with overlapping fingers (2→3, 5→4), low-set ears, micrognathia, prominent occiput, VSD, horseshoe kidney. 90% die by 1yr. Trisomy 13 (Patau): 1:15000. Cleft lip/palate, polydactyly, holoprosencephaly, microphthalmia/coloboma, scalp defects (cutis aplasia), VSD/PDA, cystic kidneys. 95% die by 1yr. Triploidy: 69 chromosomes. Partial molar pregnancy (69,XXY from dispermy). Severe IUGR, syndactyly, molar placenta. Lethal.

Turner syndrome (45,X): Only viable monosomy. 1:2500 females. Features: short stature, webbed neck, low posterior hairline, shield chest, widely spaced nipples, cubitus valgus, lymphedema (hands/feet), coarctation of aorta, bicuspid aortic valve, horseshoe kidney, streak gonads (gonadal dysgenesis → primary amenorrhea, infertility, absent puberty). Intelligence normal but nonverbal learning deficits. Mosaic forms (45,X/46,XX) can be milder. Treatment: GH for height, estrogen replacement for puberty, monitor for aortic dilation. Klinefelter syndrome (47,XXY): 1:600 males. Tall stature, gynecomastia, small firm testes, eunuchoid habitus, infertility (azoospermia), low testosterone, increased risk of breast cancer, osteoporosis, varicose veins, learning disabilities (verbal IQ < performance IQ). 80% due to maternal nondisjunction. Treatment: testosterone replacement. 47,XYY: 1:1000 males. Tall stature, normal fertility, normal intelligence, slightly increased risk of behavioral/learning difficulties (ADD, autism). 47,XXX (Triple X): 1:1000 females. Tall stature, normal phenotype, normal fertility, slightly increased risk of learning disabilities. Most undiagnosed.

22q11.2 deletion (DiGeorge/Velocardiofacial): Most common microdeletion (1:4000). Conotruncal heart defects (tetralogy of Fallot, truncus arteriosus, interrupted aortic arch type B), hypocalcemia (parathyroid aplasia), T-cell deficiency (thymic aplasia, recurrent infections, autoimmune disease), cleft palate/speech delay, dysmorphic facies (narrow palpebral fissures, tubular nose), learning disabilities, psychiatric disorders (schizophrenia risk 25x). TBX1 gene is critical. Williams syndrome (7q11.23 deletion): Elfin facies (periorbital fullness, stellate iris, full lips), supravalvular aortic stenosis (ELN gene), hypercalcemia (infancy), cocktail party personality (friendly, loquacious), intellectual disability, musculoskeletal issues, growth retardation. Prader-Willi (15q11-q13 paternal deletion): Infantile hypotonia/poor feeding, then hyperphagia/obesity (hypothalamic dysfunction), small hands/feet, almond-shaped eyes, intellectual disability, obsessive-compulsive behaviors, growth hormone deficiency. Angelman (15q11-q13 maternal deletion): Happy puppet: severe intellectual disability, jerky ataxic gait, frequent laughter/smiling, seizures, microcephaly, absent speech. UBE3A gene (expressed only from maternal allele in brain). Cri-du-chat (5p deletion): Cat-like cry (laryngeal malformation), microcephaly, hypertelorism, intellectual disability, round face. Miller-Dieker (17p13.3 deletion, LIS1): Lissencephaly (smooth brain), severe intellectual disability, seizures, dysmorphic facies. Smith-Magenis (17p11.2 deletion): Self-injurious behaviors, sleep disturbance, hoarse voice, intellectual disability, brachycephaly. Wolf-Hirschhorn (4p16.3 deletion): Greek warrior helmet facies (prominent glabella, hypertelorism), microcephaly, seizures, severe ID, cleft lip/palate. NF1 microdeletion (17q11.2): More severe neurofibromatosis type 1 with increased tumor burden, dysmorphic features, intellectual disability.

FMR1 gene CGG trinucleotide repeat expansion on X chromosome. Most common inherited cause of intellectual disability. Prevalence: 1:4000 males, 1:8000 females. Full mutation (>200 CGG repeats): Methylation of FMR1 promoter → transcriptional silencing → loss of FMRP (RNA-binding protein regulating translation at synapses). Premutation (55-200 repeats): Unstable, can expand to full mutation in offspring (anticipation, maternal transmission only). Premutation carriers at risk for: fragile X-associated tremor/ataxia syndrome (FXTAS, >50y males, intention tremor, ataxia, cognitive decline) and fragile X-associated primary ovarian insufficiency (FXPOI, 20% of carriers). Clinical features (full mutation males): Long narrow face, prominent forehead, large everted ears, macroorchidism (post-pubertal), hyperextensible joints, mitral valve prolapse, intellectual disability (moderate-severe), autism spectrum disorder (30-50%), seizures (20%), ADHD, social anxiety, hand flapping, gaze aversion. Females with full mutation: milder phenotype (skewed X-inactivation). Diagnosis: FMR1 CGG repeat analysis (PCR + Southern blot). Treatment: Symptomatic (behavioral therapy, psychostimulants, SSRIs, targeted therapies: mGluR5 antagonists, GABA agonists in trials).

Only one mutant allele needed for disease expression. Vertically transmitted, affects both sexes equally, male-to-male transmission possible. Heterozygotes affected (often more severe in homozygotes, usually embryonic lethal). Key features: Multiple generations affected, ~50% risk to offspring, variable expressivity, incomplete penetrance (some individuals with genotype do not manifest disease), de novo mutations common (e.g., NF1 ~50% de novo, achondroplasia ~80% from advanced paternal age). Haploinsufficiency: One functional copy insufficient for normal function (e.g., TBX1 in DiGeorge, ELN in Williams). Dominant negative: Mutant protein interferes with wild-type protein function (e.g., COL1A1/2 in OI, FBN1 in Marfan). Gain-of-function: Mutant protein acquires novel toxic activity (e.g., FGFR3 in achondroplasia, HTT in Huntington, KRAS in Noonan). Examples: Huntington disease, Marfan, NF1/2, Familial hypercholesterolemia (LDLR), ADPKD (PKD1/2), HHT (ENG), Ehlers-Danlos vascular (COL3A1), OI, Achondroplasia, Myotonic dystrophy, VHL, MEN1/2, TSC1/2, BRCA1/2, Lynch, FAP (APC).

Two mutant alleles required. Both sexes equally affected. Often seen in consanguineous families. Heterozygous carriers are asymptomatic. More severe than AD (loss-of-function of critical enzymes). Key features: Usually one generation affected (siblings), horizontal pattern. 25% risk to offspring of carrier parents, 50% carriers, 25% unaffected. Carrier frequency increased in certain populations (founder effect). Examples: CF (CFTR), Sickle cell (HBB), PKU (PAH), Tay-Sachs (HEXA), Gaucher (GBA), Niemann-Pick (SMPD1), GSD I (G6PC), Wilson (ATP7B), Hemochromatosis (HFE), Alpha-1 antitrypsin (SERPINA1), Friedreich ataxia (FXN), SMA (SMN1), Albinism (TYR, OCA2), Usher syndrome, Bardet-Biedl, Primary ciliary dyskinesia. Consanguinity: Increased coefficient of inbreeding (F). First cousin marriage (F=1/16): 4-6% risk of recessive disorder vs 2-3% background. Coefficient of relationship: first cousins share 1/8 of genes. Compound heterozygote: Two different mutant alleles at same locus (e.g., CF, hemochromatosis). Locus heterogeneity: Mutations in different genes cause same phenotype (e.g., OI: COL1A1/COL1A2; hearing loss: many genes).

X-Linked Recessive: Affected males (hemizygous), carrier females (usually asymptomatic). No male-to-male transmission. All daughters of affected male are obligate carriers. 50% of sons of carrier female affected. Examples: Duchenne & Becker MD (DMD), Hemophilia A (F8) & B (F9), G6PD deficiency, Fabry (GLA), Hunter/MPS II (IDS), Lesch-Nyhan (HPRT1), Color blindness, X-linked agammaglobulinemia (BTK), Wiskott-Aldrich (WAS), X-ALD (ABCD1). Duchenne MD: DMD (dystrophin), frameshift deletion/nonsense, absent protein. Becker: in-frame deletion, reduced. Onset 3-5y, Gowers sign, pseudohypertrophy of calves, waddling gait, cardiomyopathy, wheel chair by 12, death 20s (respiratory/cardiac). Creatine kinase markedly elevated. Hemophilia A: F8 intron 22 inversion common. Severity correlates with factor level. X-Linked Dominant: Both sexes affected, more severe/fatal in males (often male lethal). All daughters of affected male affected, no sons affected. Examples: Rett syndrome (MECP2, predominantly females due to male lethality), Incontinentia pigmenti (IKBKG/NEMO, male lethal), Vitamin D-resistant rickets/hypophosphatemic rickets (PHEX), Goltz syndrome (PORCN, male lethal), Alport syndrome (COL4A5, also AD/AR forms). X-inactivation (Lyonization): Random inactivation of one X in each cell in early development. Carrier females may have mild expression of X-linked recessive disorders (skewed inactivation). Mosaic expression (e.g., Rett, Fabry carriers with symptoms).

Maternal transmission only. mtDNA: 16.5kb circular, 37 genes (13 proteins, 22 tRNAs, 2 rRNAs). High mutation rate (lack of histones, limited repair). Heteroplasmy: variable proportion of mutant mtDNA within cells. Threshold effect: clinical expression when mutant proportion exceeds tissue-specific threshold. Mitotic segregation. Examples: LHON (ND1/4/6, sudden vision loss, young men), MELAS (MT-TL1, encephalopathy, lactic acidosis, stroke-like episodes), MERRF (MT-TK, myoclonic epilepsy, ragged red fibers), Kearns-Sayre (deletions, progressive external ophthalmoplegia, heart block, ataxia), Leigh syndrome (complex I/IV genes, subacute necrotizing encephalomyelopathy), Pearson syndrome (mtDNA deletion, sideroblastic anemia + pancreatic insufficiency), NARP (MT-ATP6, neuropathy, ataxia, retinitis pigmentosa). Diagnosis: Muscle biopsy (ragged red fibers, COX-deficient fibers), mtDNA sequencing. Recurrence risk: ~100% if mother has mutation (but variable heteroplasmy, uncertain phenotype). No transmission from affected male.

Anticipation: Earlier onset/increased severity in successive generations due to unstable trinucleotide repeat expansions. Polyglutamine (CAG) disorders: CAG repeat in coding region → expanded polyglutamine tract → protein aggregation, toxic gain-of-function. Anticipation more pronounced with paternal transmission. Examples: Huntington disease (HTT, 4p16.3, normal <26, penetrant >40, CAG), SBMA/Kennedy disease (AR), SCAs 1/2/3/6/7/17, DRPLA. Non-coding repeat expansions: Fragile X (FMR1 CGG 5'UTR, >200 full, methylation silencing). Myotonic dystrophy type 1 (DMPK CTG 3'UTR, 50-4000, RNA toxicity, MBNL splicing dysregulation). Friedreich ataxia (FXN GAA intron 1, 70-1700, transcriptional silencing, frataxin deficiency). DM2 (CNBP CCTG intron 1, 75-11000, milder, proximal weakness). C9orf72 (GGGGCC intron 1, 30-2000, ALS + FTD, both gain + loss of function). Parental bias: HD & SCA: paternal → larger expansions. DM1 & Fragile X: maternal → larger expansions. FRDA: paternal contractions, maternal expansions.

Imprinting: Epigenetic phenomenon where genes are expressed only from one parental allele. Imprinting control regions (ICRs) differentially methylated. Prader-Willi (PWS): 15q11-q13, loss of paternally expressed genes (SNRPN, NDN, MKRN3, MAGEL2). 70% de novo paternal deletion, 25% maternal UPD, <5% imprinting defect. Angelman (AS): Same region, loss of maternally expressed UBE3A. 70% de novo maternal deletion, 2-5% paternal UPD, 3-5% imprinting defect, 10% UBE3A mutation. Beckwith-Wiedemann (BWS): 11p15.5, overgrowth, macrosomia, omphalocele, macroglossia, hemihypertrophy, embryonal tumors (Wilms, hepatoblastoma). ICR1 (IGF2/H19) hypermethylation, ICR2 (KCNQ1OT1/CDKN1C) hypomethylation. Russell-Silver (SRS): 11p15.5 hypomethylation (50%) or maternal UPD 7 (10%). IUGR, triangular face, body asymmetry. TNDM: 6q24, paternal UPD or duplication. UPD testing: SNP microarray or methylation-specific MLPA. Recurrence risk low for UPD but may recur with certain translocations.

Gonadal mosaicism: Mutation present in some germ cells but not detectable in blood. Important for recurrence risk when disease appears de novo. Example: DMD, OI, tuberous sclerosis, NF1. Recurrence risk for sibs: up to 10-15% for certain disorders even if parents unaffected. Empiric risk: ~1-2% for gonadal mosaicism after de novo AD mutation. Multifactorial inheritance: Genetic (polygenic) + environmental. Examples: NTD, cleft lip/palate, CHD, hypertension, diabetes, autism. Recurrence risk: 2-5% for first-degree relatives (higher if severe, multiple affected, same sex for sex-limited traits). Threshold model: continuous liability distribution with threshold for disease expression. Somatic mosaicism: Post-zygotic mutation in subset of cells. Examples: McCune-Albright (GNAS, polyostotic fibrous dysplasia, cafe-au-lait, endocrine overactivity), Sturge-Weber (GNAQ, port-wine stain), Proteus (AKT1), CLOVES (PIK3CA), segmental NF1. Heteroplasmy: Mitochondrial mosaicism.

Principles: Allele and genotype frequencies remain constant across generations in the absence of evolutionary forces. Equations: p + q = 1 (allele frequencies). p² + 2pq + q² = 1 (genotype frequencies). p = frequency of dominant/normal allele (A), q = frequency of recessive/mutant allele (a). Assumptions: Random mating, no mutation, no selection, no migration, infinite population size, discrete generations, equal allele frequencies in sexes. Deviations indicate evolutionary forces. Carrier frequency calculation (AR disease): Incidence = q². For 1/10,000: q² = 0.0001, q = 0.01, p = 0.99, carrier frequency 2pq = 0.0198 (~1/50). For 1/40,000: q = 0.005, p = 0.995, carrier ~1/100. X-linked recessive: Disease frequency in males = q. Carrier frequency in females = 2pq. Incidence in females = q² (rare). Multiple alleles: (p + q + r)² = 1 (ABO, HLA). Testing for HWE: Chi-square test comparing observed vs expected genotype counts. Deviation may indicate genotyping error, population stratification, or selection.

Fitness (w): Relative reproductive success. Selection coefficient (s = 1 - w). Against recessive homozygotes: q decreases slowly (most deleterious alleles hidden in heterozygotes). For rare recessive lethal (q is small, e.g., 0.01), selection is ineffective. Against dominant alleles: Strong selection (all carriers affected). Most AD diseases persist due to de novo mutations (achondroplasia s ~0.8). Heterozygote advantage: Heterozygote has higher fitness than either homozygote. Classic: sickle cell trait (HbAS) confers malaria resistance (w ~1.2-1.3 in endemic regions). Balanced polymorphism maintains high HbS frequency (up to 20% in parts of Africa). Other: G6PD deficiency, thalassemias, CFTR (possible typhoid resistance), CCR5-Delta32 (HIV resistance). Frequency-dependent selection: Rare alleles have advantage (MHC diversity). Mutation-selection balance: For AR: q = √(μ/s). For AD: q = μ/s. Used to estimate mutation rates for lethal disorders.

Genetic drift: Random fluctuations in allele frequencies in small populations. Leads to fixation or loss. Founder effect: Small number of founders establish new population, specific alleles become enriched. Ashkenazi Jewish founder mutations: Tay-Sachs (HEXA 1278insTATC, 1:25 carrier), Gaucher (GBA N370S, 1:15), Canavan (ASPA E285A, 1:40), familial dysautonomia (IKBKAP, 1:30), Bloom (BLM, 1:100), Niemann-Pick A (SMPD1, 1:90), Fanconi C (FANCC, 1:100), Factor XI (F11, 1:10). Finnish: Congenital nephrotic (NPHS1), aspartylglucosaminuria (AGA), diastrophic dysplasia (SLC26A2). French Canadian: Tay-Sachs, CF (444delA), LPL deficiency, OPMD (PABPN1). Amish/Mennonite: MSUD (BCKDHA), Glutaric acidemia I (GCDH), Ellis-van Creveld (EVC/EVC2), cartilage-hair hypoplasia (RMRP). Bottleneck: Drastic population reduction → loss of diversity. Out-of-Africa bottleneck → lower diversity in non-Africans. Effective population size (N_e): Size of ideal population with same drift rate, usually much smaller than census size.

Linkage disequilibrium (LD): Non-random association of alleles at different loci. D' and r² measures. LD blocks separated by recombination hotspots. GWAS: Tests millions of common SNPs (MAF >5%) for association with traits/diseases. Case-control or quantitative trait. Uses imputation (1000 Genomes, TOPMed). Population structure correction (PCA, mixed models). Multiple testing correction (p < 5 × 10^{-8}). Findings: Most variants have small effect sizes (OR 1.1-1.5). Explain small fraction of heritability (missing heritability). Over 5000 GWAS loci identified. Post-GWAS: Fine-mapping (credible sets), functional annotation (eQTL, pQTL, splicing QTL), colocalization, Mendelian randomization. Polygenic risk scores (PRS): Weighted sum of risk alleles across many SNPs. Methods: LDpred, PRS-CS, Bayesian regression. Used for risk stratification (CAD, breast cancer, T2D). Clinical utility still limited. Heritability: Proportion of phenotypic variance due to genetics. Broad-sense (H², all genetic) vs narrow-sense (h², additive). Twin studies (MZ vs DZ), family studies, SNP-based heritability (GREML, LDSC). h² height ~0.8, BMI ~0.4-0.7, schizophrenia ~0.8, autism ~0.8, breast cancer ~0.3. Missing heritability: Gap between GWAS and family-based heritability. Sources: rare variants, structural variants, epistasis, GxE, epigenetic effects.

Karyotype (G-banding): Gold standard for chromosome abnormalities. Requires metaphase cells. Resolution: 400-550 bands, detects abnormalities >5-10 Mb. Indications: suspected aneuploidy, structural rearrangements, hematologic malignancies, reproductive genetics. FISH: Targeted DNA probes, metaphase or interphase. Resolution: specific locus. Uses: rapid aneuploidy detection (chr13/18/21/X/Y), microdeletion syndromes (DiGeorge, Williams, Cri-du-chat), HER2 amplification, BCR-ABL fusion, MLL rearrangements. Chromosomal microarray (CMA): First-line for unexplained DD/ID, autism, MCA. Detects CNVs at 50-100 kb resolution. Array CGH (comparative genomic hybridization) or SNP array (detects UPD, consanguinity, mosaicism). Diagnostic yield ~15-20% in DD/ID (vs ~3% karyotype). Limitations: balanced rearrangements, low-level mosaicism, point mutations. ACMG/ISCA guidelines: report CNVs >400-500 kb, evaluate for known syndromes.

Sanger sequencing: Gold standard for single-gene testing. Dideoxy chain termination. Read length ~700-900bp. Used for confirmation of NGS variants, single-gene disorders (CFTR, FBN1, BRCA1/2 germline). Next-generation sequencing (NGS): Massively parallel. Platforms: Illumina (sequencing by synthesis, short reads 150-300bp), Ion Torrent (semiconductor), PacBio HiFi (long reads 10-25kb), Oxford Nanopore (very long reads). Targeted gene panels: 5-500 genes, high depth (100-1000x), fewer VUS. Applications: hereditary cancer, cardiomyopathy, epilepsy, hearing loss, RASopathy panels. Whole exome sequencing (WES): Captures all coding exons (~1% genome, ~20,000 genes). ~60-80x depth. Diagnostic yield ~25-40% in selected cases (DD/ID, MCA, neurological). Trio WES improves yield. Whole genome sequencing (WGS): Covers coding + non-coding. Higher cost, more incidental findings. Emerging as first-line. Long-read sequencing: Solves SVs, repeats, phasing, segmental duplications. Used for repeat expansions (Fragile X, HD, DM1, C9orf72), HLA typing. RNA sequencing: Detects aberrant splicing, allelic imbalance, fusions, expression outliers. Adds 10-15% diagnostic yield when added to WES/WGS.

ACMG/AMP 2015: Five tiers: Pathogenic (P), Likely Pathogenic (LP), VUS, Likely Benign (LB), Benign (B). Pathogenic evidence: PVS1 (null variant in LOF gene - nonsense, frameshift, canonical splice, initiation codon). PS1 (same amino acid change as known P). PS2 (de novo confirmed). PS3 (functional assay damaging). PS4 (prevalence in affected increased). PM1 (hotspot/functional domain). PM2 (absent in gnomAD). PM3 (in trans with P for recessive). PM4 (protein length change). PM5 (novel missense at same codon). PP1 (cosegregation). PP2 (missense in gene with few benign). PP3 (computational tools predict damaging). PP4 (highly specific phenotype). Benign evidence: BA1 (MAF >5%). BS1 (MAF >expected). BS2 (observed in healthy adult). BS3 (functional benign). BP1-7 (various). Combination rules: P: 1 PVS1 + >=1 PS/PM/PP, or >=2 PS, or 1 PS + >=3 PM, or 1 PS + 2 PM + >=2 PP, or >=3 PM + >=2 PP. VUS: Insufficient/conflicting evidence. Up to 30% of WES findings. May be reclassified. Recontact policy critical. Splice prediction: SpliceAI (deep learning), MaxEntScan. Population databases: gnomAD (~76k genomes, ~126k exomes). ClinVar, LOVD, HGMD, ClinGen. ClinGen gene-disease validity: Definitive, strong, moderate, limited, disputed, refuted.

NIPT: Cell-free fetal DNA from maternal blood, >10 weeks. Detection >99% T21, ~97% T18, ~92% T13. Also sex aneuploidies, selected microdeletions (22q11.2, 1p36, Angelman, Prader-Willi, Cri-du-chat). Screening test only - confirm with CVS/amnio. CVS: 10+0 to 13+6 weeks. Transcervical or transabdominal. Miscarriage risk ~0.2%. Confined placental mosaicism risk ~1%. Amniocentesis: 15+0 to 20+6 weeks. Miscarriage risk ~0.1-0.2%. Preferred after 15 weeks. PGT: IVF embryo biopsy day 5/6 (trophectoderm). PGT-M (monogenic), PGT-A (aneuploidy), PGT-SR (structural). Requires probe development for PGT-M. Carrier screening: ACOG recommends CF (pan-ethnic), SMA (pan-ethnic), hemoglobinopathies (ethnic), fragile X (if FH). Expanded carrier screening (274+ genes). AJ panel: Tay-Sachs, Canavan, familial dysautonomia, Bloom, Fanconi C, Niemann-Pick A, mucolipidosis IV. Newborn screening (NBS): Dried blood spot 24-48h. RUSP: 35+ core conditions (PKU, MSUD, MCAD, GA1, CH, CAH, SCD, CF, hearing loss, critical CHD). Tandem MS. Molecular confirmatory testing (CFTR, ACADM). DTC testing: 23andMe FDA-authorized for limited BRCA1/2 AJ variants, MUTYH, CF, Bloom, G6PD, Factor XI, HH. Clinical confirmation required. Not medical grade.

Pedigree symbols: Square (male), circle (female), filled (affected), diagonal (deceased), arrow (proband). Consanguinity: double line. Bayesian calculation: Prior × conditional = joint, then normalize. Refines risk with incomplete penetrance or test results. Example: BRCA1/2, unaffected 45yo woman, affected mother/sister (known familial mutation). Prior: 50% inherited mutation (AD). Conditional: unaffected at 45. BRCA1 penetrance by 45 ~45%. Conditional no phenotype given mutation = 0.55. Conditional no phenotype without mutation = ~1. Joint mutation = 0.5 × 0.55 = 0.275. Joint no mutation = 0.5 × 1 = 0.5. Posterior = 0.275/(0.275+0.5) = 0.355 (35.5%). Counselling framework: Pre-test: purpose, outcomes (positive/negative/VUS/incidental), family implications, GINA. Post-test: results, medical management, family communication, psychological support, reproductive options.

Oncogenes: Gain-of-function, dominant. Normal proto-oncogenes regulate cell growth/survival. Activated by mutation, amplification, translocation, fusion. Examples: RAS (KRAS/NRAS/HRAS, ~30% of cancers, G12C/D/V, GTPase locked active). MYC (amplified in many cancers). EGFR (TKI-sensitive L858R/exon19 del, resistant T790M/C797S). HER2 (amplified breast/gastric). BRAF V600E (melanoma/colon/thyroid). BCR-ABL (t(9;22) CML). ALK fusions (EML4-ALK NSCLC). NTRK fusions (tumor-agnostic). IDH1/2 (glioma/AML). Tumor suppressor genes (TSGs): Loss-of-function, recessive. Knudson two-hit: both alleles inactivated. First hit germline (hereditary) or somatic. Second hit: LOH, deletion, mutation, promoter hypermethylation. Gatekeeper TSGs: Directly regulate growth (RB1, APC, PTEN, NF1). Caretaker TSGs: Maintain genome stability (BRCA1/2, MSH2/MLH1, ATM, BLM, WRN). Examples: TP53 (Li-Fraumeni, most mutated gene in cancer). RB1 (retinoblastoma, cell cycle). APC (FAP, WNT pathway). BRCA1/2 (HBOC, HR repair). VHL (HIF, RCC). PTEN (Cowden, PI3K/AKT). NF1 (RAS-GAP, neurofibromas). CDKN2A/p16 (melanoma/pancreas). Synthetic lethality: PARP inhibition in BRCA-deficient tumors (SSB repair impaired → DSB, HR deficient → cell death).

MMR (Mismatch Repair): Corrects replication errors. MLH1/MSH2/MSH6/PMS2. Defect → MSI-H, hypermutation. Lynch syndrome: MMR germline mutations. Colon 50-80% (right-sided, synchronous), endometrial 40-60%, ovarian 10-15%, also gastric, small bowel, ureter, biliary, pancreas, brain (Turcot), sebaceous (Muir-Torre). Amsterdam II criteria, Bethesda guidelines. IHC for MMR loss, MSI testing. Aspirin reduces CRC (CAPP2). NER (Nucleotide Excision Repair): Repairs bulky DNA adducts, UV dimers. XP proteins. Defect → xeroderma pigmentosum (1000x skin cancer, neurologic). Cockayne syndrome (photosensitivity, growth failure, no increased cancer). Trichothiodystrophy (brittle hair, ichthyosis). HR (Homologous Recombination): Error-free DSB repair. BRCA1/2, PALB2, RAD51, ATM, ATR, NBS1, MRE11. HBOC: BRCA1 (45-65% breast, 15-40% ovarian, triple-negative). BRCA2 (45-55% breast, 10-20% ovarian, male breast ~7%). PALB2 (~30% breast). RRSO by 35-40, risk-reducing mastectomy, enhanced screening (MRI + mammo). PARP inhibitors. Platinum sensitivity. Li-Fraumeni: TP53. Sarcomas, breast, brain, adrenocortical, leukemia, early onset, multiple primaries. Avoid radiation. FAP: APC. Hundreds of polyps by teens, 100% colon cancer by 40. Gardner (osteomas/desmoids/CHRPE). Turcot (brain). Cowden: PTEN. Hamartomas, high breast/thyroid/endometrial, macrocephaly, trichilemmomas. Peutz-Jeghers: STK11/LKB1. Hamartomatous polyps, mucocutaneous pigmentation, high GI/breast/ovary/pancreas/cervix. NF1: Neurofibromas, cafe-au-lait, optic gliomas, MPNST, pheochromocytoma. NF2: Schwannomas, meningiomas, ependymomas. TSC1/2: Hamartomas in brain/lung/kidney/skin, SEGA, AML, LAM, epilepsy. VHL: Hemangioblastoma, RCC, pheochromocytoma, PNET. MEN1/2: Parathyroid/pituitary/pancreas (MEN1). MTC/pheochromocytoma/hyperparathyroidism (MEN2, RET). RB1: Retinoblastoma, trilateral. WT1: Wilms tumor. Gorlin/BCNS: PTCH1. Basal cell carcinomas, jaw keratocysts, skeletal.

Driver vs passenger: Driver confers selective advantage. Most tumors have 2-10 drivers. TMB: Somatic mutations/Mb. High TMB (>10 mut/Mb) → immunotherapy response. Pembrolizumab approved tumor-agnostic for TMB-H. Mutational signatures (COSMIC): SBS1 (deamination), SBS2/13 (APOBEC), SBS4 (tobacco), SBS7 (UV), SBS10 (POLE), SBS15/20 (MMR deficiency), SBS3 (HRD/BRCA). Chromothripsis: Catastrophic shattering of chromosomes. Common in osteosarcoma. Kataegis: Localized hypermutation (APOBEC). Tumor heterogeneity: Intratumoral, branched evolution, subclonal drivers. Liquid biopsy: ctDNA for early detection, MRD, monitoring, resistance mutations. Guardant360 CDx, FoundationOne Liquid CDx. Tumor-agnostic approvals: Pembrolizumab (MSI-H, TMB-H). Larotrectinib/entrectinib (NTRK fusions). Dostarlimab (dMMR). Selpercatinib (RET). Immunogenomics: Neoantigens, TILs, PD-L1, interferon gamma signature. Whole genome doubling: Common (30-50%), prognostic and therapeutic implications.

CYP2D6: Highly polymorphic, >100 star alleles. Metabolizes ~25% of drugs (codeine, tramadol, tamoxifen, TCAs, SSRIs, beta-blockers, antipsychotics). Phenotypes: PM (~7-10% Europeans, ~2% Asians), IM, NM, UM (~1-2% Europeans, up to 30% Ethiopian/Saudi). Codeine: Prodrug activated by CYP2D6 to morphine. UM → respiratory depression. FDA black box: contraindicated children <12, post-tonsillectomy. Tamoxifen: Requires CYP2D6 for endoxifen. PM → reduced efficacy → higher recurrence. TCAs: PM → toxicity (QT prolongation). UM → no efficacy. CYP2C19: Clopidogrel (prodrug). PM (~2-5% Europeans, ~15% Asians) → reduced activation → stent thrombosis/MI. FDA black box. Alternative: prasugrel/ticagrelor. PPIs: PM → higher efficacy. Citalopram/escitalopram: PM → QT risk (max 20mg). CYP2C9: Warfarin (S-warfarin). PM (*2/*3) → reduced clearance, bleeding risk. Reduced starting dose. CYP3A4/5: Most abundant CYP. Tacrolimus dosing based on CYP3A5 (expressor vs non-expressor). CYP2B6: Efavirenz PM → CNS toxicity. CYP4F2: Warfarin, V433M increases dose requirement.

TPMT: Thiopurine metabolism (azathioprine, 6-MP, thioguanine). PM (1:300, TPMT*2/*3A/*3B/*3C) → severe myelosuppression. Heterozygotes (10%) → dose reduce 30-50%. Pre-test standard. NUDT15: Additional risk in Asians (p.Arg139Cys, ~10-15%). DPYD: Rate-limiting enzyme for 5-FU catabolism. Reduced function (*2A, c.2846A>T, c.1679T>G, HapB3) → severe toxicity (mucositis, diarrhea, neutropenia, HFS, death). 3-5% population carriers. Dose reduce 50% for heterozygotes, avoid for homozygotes. Pre-treatment uracil screening. UGT1A1: Glucuronidates bilirubin and drugs. UGT1A1*28 (Gilbert, ~10% homozygotes). Irinotecan: SN-38 active metabolite. *28 → increased SN-38 → severe diarrhea/neutropenia. FDA label: reduce dose. G6PD: X-linked, protects RBCs from oxidative stress. Class I-V. African variant G6PD A- (~10-15% AA men, Class III). Triggers: dapsone, primaquine, sulfonamides, nitrofurantoin, rasburicase, fava beans, infections. Hemolysis, jaundice, dark urine. Screen before primaquine. ALDH2: Asian variant (*2, ~40% East Asians) → flushing, nausea, increased esophageal cancer risk.

HLA-B*5701: Abacavir hypersensitivity (fever, rash, GI/respiratory, hypotension, death on rechallenge). Mandatory screening. Positive = never use. Prevalence: 5-8% Europeans, ~1% African, <1% East Asian. HLA-B*1502: Carbamazepine SJS/TEN in Asians. Screen in at-risk (Han Chinese, Thai, Malay, Indonesian, Filipino, Vietnamese). Avoid carbamazepine (also oxcarbazepine, phenytoin, lamotrigine; cross-reactivity). HLA-B*5801: Allopurinol SJS/TEN in Han Chinese, African American, European. Screen high-risk. Alternative: febuxostat. HLA-A*3101: Carbamazepine hypersensitivity (MPE, DRESS, SJS/TEN) in Europeans/Japanese. Mechanism: Abacavir binds HLA-B*57:01 F pocket, alters peptide repertoire, triggers CD8+ T-cell response. Not dose-dependent. CPIC levels: Mandatory (abacavir), strong (carbamazepine), moderate (allopurinol). One-time test, integration into EHR.

Warfarin: S-warfarin (active) by CYP2C9. Inhibits VKORC1. CYP2C9 reduced (*2/*3): 25-75% lower dose. VKORC1 (-1639G>A): AA → ~50% lower dose. GG ~2x dose. Frequency: ~40% East Asian, ~15% European, ~5% African. CYP4F2 V433M: ~10% higher dose. Algorithm (IWPC): Includes CYP2C9, VKORC1, age, height, weight, race, inducers, amiodarone. FDA PGx dosing tables. SLCO1B1 (OATP1B1): Hepatic statin uptake. *5 (c.521T>C) → increased simvastatin → myopathy (OR 4.5 heterozygote, 17 homozygote). CPIC: avoid simvastatin 80mg in heterozygotes. Atorvastatin/rosuvastatin also affected. ABCB1 (P-gp): Digoxin, tacrolimus, cyclosporine. C3435T variant. Clinical utility limited. CFTR modulators: Precision therapy for CF. Ivacaftor (G551D). Lumacaftor/ivacaftor (F508del). Elexacaftor/tezacaftor/ivacaftor/Trikafta (F508del, ~90% of CF).

EGFR: NSCLC L858R/ex19del → osimertinib (first-line). T790M → osimertinib. C797S → pan-resistance. BRAF V600E: Melanoma, NSCLC, CRC, thyroid, HCL. Dabrafenib + trametinib (first-line). Encorafenib + binimetinib. BCR-ABL: CML, ALL. Imatinib (first-line). Dasatinib, nilotinib, bosutinib, ponatinib. T315I: ponatinib/asciminib. HER2: Breast, gastric. Trastuzumab, pertuzumab, T-DM1, T-DXd, lapatinib, neratinib, tucatinib. ALK: NSCLC (~5%). Crizotinib, alectinib, brigatinib, ceritinib, lorlatinib. NTRK: Tumor-agnostic. Larotrectinib, entrectinib (high CNS). IDH1/2: AML, glioma, cholangiocarcinoma. Ivosidenib, enasidenib, vorasidenib (glioma). RET: NSCLC, medullary thyroid. Selpercatinib, pralsetinib. METex14: NSCLC. Capmatinib, tepotinib. FGFR: Bladder (FGFR3), cholangiocarcinoma (FGFR2). Erdafitinib, pemigatinib, infigratinib. HRD: PARPi (olaparib, niraparib, rucaparib, talazoparib). BRCA+ breast/ovarian/pancreas/prostate. Immune checkpoint: PD-L1, MSI-H, TMB-H. Pembrolizumab, nivolumab, atezolizumab, durvalumab, ipilimumab. Gene fusions: BCR-ABL, EML4-ALK, TMPRSS2-ERG (prostate), EWSR1-FLI1 (Ewing), SS18-SSX (synovial), FUS-DDIT3 (myxoid liposarcoma), PAX3/7-FOXO1 (alveolar RMS). RNA-seq for detection.

Key periods: Embryonic (3-8 weeks): organogenesis, most sensitive to teratogens. Fetal (9 weeks-term): growth/maturation. Neural crest: Multipotent cells from neural tube. Derivatives: peripheral NS, melanocytes, adrenal medulla, craniofacial mesenchyme, cardiac outflow tract, parafollicular C-cells, Schwann cells. Gene regulatory networks: HOX (anterior-posterior patterning, clustered 4 groups A-D, collinear). PAX (CNS, eye, kidney). SOX (sex determination, neural, chondrogenesis). SHH (ventral neural tube, limb, craniofacial). WNT (β-catenin canonical, planar cell polarity). FGF (patterning, limb, craniofacial). TGF-β/BMP (DV patterning, bone, left-right via NODAL). NOTCH (lateral inhibition, neurogenesis, somitogenesis). Left-right asymmetry: NODAL/LEFTY/PITX2. Defects → situs inversus, heterotaxy (Ivemark). Kartagener (primary ciliary dyskinesia, DNAI1/DNAH5, 50% situs inversus). Somite formation: Segmentation clock (NOTCH/WNT/FGF). Sclerotome (vertebrae), dermomyotome (muscle/dermis). Defects → Klippel-Feil, spondylocostal dysostosis.

Neurulation: Primary (days 18-26, brain/spinal cord to S2). Secondary (days 26-28, sacral cord). NTDs from failed closure. Types: Anencephaly (anterior neuropore, lethal). Spina bifida: myelomeningocele (cord + meninges), meningocele (meninges only), occulta (bony defect, asymptomatic). Encephalocele (skull protrusion). Etiology: Multifactorial. MTHFR C677T, folate deficiency, valproate (10x), carbamazepine, methotrexate. Prevention: Folic acid 0.4-0.8mg daily all women. High-risk: 4-5mg daily, 3mo preconception. Screening: MSAFP 15-20wk (elevated 2.5x open NTDs). US anatomy scan. Amniotic AFP + AChE. Recurrence: 2-5% after one affected. Treatment: Surgical closure 24-48h, VP shunt, multidisciplinary. Fetal surgery (MOMS trial: prenatal closure reduces shunting, improves motor). Syndromic NTDs: Meckel-Gruber (encephalocele, polydactyly, renal cysts, lethal), Walker-Warburg (cobblestone lissencephaly), Jarcho-Levin (spondylocostal).

Cleft lip/palate: Multifactorial 1:1000. Fusion failure 4-7wk (CL) / 8-12wk (CP). Recurrence ~4%. Syndromic: Van der Woude (IRF6, lower lip pits, AD, most common syndromic CL/P). Pierre Robin (micrognathia + glossoptosis + CP, can be isolated or Stickler/22q11.2). Treacher Collins (TCOF1, malar hypoplasia, micrognathia, coloboma, hearing loss). Craniosynostosis: Sagittal (scaphocephaly), coronal (brachycephaly, FGFR). Apert (FGFR2, coronal + syndactyly). Crouzon (FGFR2/3, exophthalmos, no hands). Pfeiffer (FGFR1/2, broad thumbs). Muenke (FGFR3 P250R, hearing loss). Saethre-Chotzen (TWIST1, asymmetric). Achondroplasia: FGFR3 G380R, AD, mostly de novo (advanced paternal age). Rhizomelia, macrocephaly, foramen magnum stenosis, spinal stenosis. Vosoritide. OI: COL1A1/A2 (AD, 90%). Blue sclerae, hearing loss, dentinogenesis imperfecta, fractures. Type I (mild) to II (lethal). Bisphosphonates. Marfan: FBN1 (AD, ~25% de novo). Ectopia lentis (upward), aortic root aneurysm, arachnodactyly, pectus, scoliosis, dural ectasia. Ghent criteria. Losartan/BB, elective root replacement (>50mm). Ehlers-Danlos: Classical (COL5A1/A2, hyperextensible skin, atrophic scars). Hypermobility (hEDS, unknown). Vascular (COL3A1, arterial/bowel/uterine rupture, early death, avoid elective surgery). Stickler: COL2A1 (AD). Pierre Robin, myopia/retinal detachment, hearing loss, midface hypoplasia. Loeys-Dietz: TGFBR1/2. Aggressive aortic aneurysm, arterial tortuosity, hypertelorism, bifid uvula.

CHD: ~1% of births. Recurrence 2-5%. Genetic: Trisomy 21 (AVSD), 18 (VSD), 13 (VSD/PDA), Turner (coarctation/bicuspid). TBX5 (Holt-Oram: ASD + thumb anomalies). NKX2-5 (ASD, AV block). GATA4 (ASD/VSD). NOTCH1 (bicuspid AV, LVOT, aortic aneurysm). CHD7 (CHARGE). RASopathies (Noonan syndrome): PTPN11 (50%, SHP2 GOF). SOS1, RAF1, RIT1, KRAS, NRAS, BRAF, MAP2K1/2. Features: webbed neck, pectus, PS (most common CHD), HCM, cryptorchidism, bleeding, mild ID, short stature, lymphatic. AD. LEOPARD: PTPN11/RAF1, lentigines, deafness, HCM. CFC: BRAF, severe ID, ectodermal, HCM. Costello: HRAS G12S, coarse, severe ID, HCM, cancer (rhabdomyosarcoma/bladder). CHARGE: CHD7 (AD, 65% de novo). Coloboma, Heart (conotruncal), Choanal atresia, Retarded growth, Genital hypoplasia, Ear anomalies. VACTERL: Vertebral, Anorectal, Cardiac (VSD), TE fistula, Renal, Limb (radial). At least 3.

Alcohol/FAS: Leading preventable ID. Microcephaly, short palpebral fissures, smooth philtrum, thin vermillion, ID, CHD. No safe level. Isotretinoin: Highest risk. CNS (hydrocephalus), craniofacial (microtia/anotia), CHD (conotruncal), thymic aplasia. iPledge. Valproate: 10-15% malformations (NTD ~10%, hypospadias, cleft palate, CHD). Neurodevelopmental: reduced IQ, autism 3-5x. Avoid in pregnancy. Warfarin: Embryopathy (nasal hypoplasia, stippled epiphyses). Avoid (use LMWH). ACEi/ARB: 2nd/3rd trimester: fetal renal failure, oligohydramnios, pulmonary hypoplasia, skull defects. Fetotoxic. Maternal PKU: Uncontrolled Phe → microcephaly, ID, CHD, IUGR. Preconception diet (Phe 120-360). Maternal diabetes: Sacral agenesis (200x), NTD, CHD, macrosomia. Lithium: Ebstein anomaly (1:1000 vs 1:20000). Antiepileptics: Phenytoin (fetal hydantoin: IUGR, microcephaly, ID, cleft palate). Carbamazepine (spina bifida ~1%). Topiramate (cleft lip ~2%). TORCH: Toxoplasma (hydrocephalus, chorioretinitis, intracranial calcifications). Rubella (deafness, cataracts, PDA, IUGR). CMV (most common congenital infection, hearing loss, microcephaly, periventricular calcifications, ID). HSV (vesicular, encephalitis, disseminated). Syphilis (snuffles, saddle nose, Hutchinson teeth, saber shins). Zika (microcephaly, brain calcifications). Parvovirus B19 (hydrops fetalis). Radiation: >0.1 Gy: microcephaly, ID, IUGR. Methotrexate: Aminopterin syndrome (cranial dysostosis, limb defects, ID).

PKU: PAH (AR). Phe hydroxylase deficiency → Phe accumulation. Types: classic, mild, HPA. BH4 cofactor defects. NBS. Clinical: progressive ID (if untreated), microcephaly, seizures, eczema, fair skin, musty odor. Treatment: Phe-restricted diet, Phe-free formula, sapropterin (BH4-responsive ~30%), pegvaliase (PEG-PAL). Maternal PKU: strict preconception diet (Phe 120-360). MSUD: BCKDHA/B (AR). Branched-chain ketoacid dehydrogenase deficiency. Elevated Leu/Ile/Val. MSUD odor. Encephalopathy, cerebral edema. Treatment: BCAA-free formula, thiamine (mild), dialysis (acute), liver transplant. Homocystinuria: CBS (AR). Elevated Met + Hcy. Ectopia lentis (downward/inward), marfanoid, ID, thromboembolism, osteoporosis. Treatment: B6 (50% responsive), betaine, B12/folate, low-Met diet. MMA: MUT/MMAA/BB/CHC/D (AR). Elevated C3. Metabolic acidosis, hyperammonemia, encephalopathy, neutropenia, CKD, basal ganglia damage. Treatment: protein restriction, carnitine, B12 (responsive forms), N-carbamylglutamate. Propionic acidemia: PCCA/PCCB (AR). Similar. Tyrosinemia I: FAH (AR). Succinylacetone (pathognomonic). Hepatic failure, Fanconi, porphyria-like crisis, HCC. Treatment: nitisinone (NTBC), Tyr-restricted diet, liver transplant.

Enzyme defects in ammonia to urea conversion. OTC deficiency: X-linked, most common. Variable in carrier females. CPS1, ASS (citrullinemia), ASL (argininosuccinic aciduria), ARG1 (arginase, milder, spastic diplegia): AR. Clinical: Hyperammonemia (neonatal or later): vomiting, lethargy, encephalopathy, cerebral edema, coma, death. Respiratory alkalosis. Diagnosis: Plasma NH3 (profound >200-1000+), amino acids (glutamine/alanine elevated, specific pattern), urine orotic acid (high in OTC, low in CPS). Treatment: Acute: stop protein, IV glucose + lipids, Na benzoate/Na phenylacetate IV, arginine (except ARG1), carbamylglutamate (NAGS), hemodialysis. Chronic: protein restriction, NH3 scavengers (Na phenylbutyrate/glycerol phenylbutyrate), arginine/citrulline, essential AA, liver transplant.

GSD I (von Gierke): G6PC (AR). G6Pase deficiency. Severe fasting hypoglycemia, lactic acidosis, hyperuricemia, hyperlipidemia, hepatomegaly, growth failure, bleeding, gout, renal disease, hepatic adenomas (HCC risk). Treatment: raw cornstarch q3-4h, allopurinol. GSD II (Pompe): GAA (AR). Acid alpha-glucosidase deficiency. Infantile: hypotonia, cardiomegaly, macroglossia, death <1yr. Late-onset: proximal myopathy, respiratory insufficiency. ERT (alglucosidase alfa, avalglucosidase alfa). NBS available. GSD III (Cori): AGL (AR). Debranching enzyme. Milder than I, myopathy, cardiomyopathy. GSD V (McArdle): PYGM (AR). Muscle phosphorylase. Exercise intolerance, myoglobinuria, second wind. GSD VI (Hers): PYGL (AR). Mild hypoglycemia, hepatomegaly. GSD VII (Tarui): PFKM (AR). No second wind, hemolytic anemia. GSD IX: PHKA2 (mild, ketotic hypoglycemia, improves with age). GSD IV: GBE1 (branching, severe, hepatic failure).

Gaucher: GBA (AR). Glucocerebrosidase deficiency → glucocerebroside in macrophages (Gaucher cells). Type 1 (AJ, 1:15 carrier): hepatosplenomegaly, bone pain/crisis, thrombocytopenia. Type 2 (infantile, neurologic, death <2yr). Type 3 (chronic neuronopathic). Diagnosis: enzyme activity, GBA sequencing. Treatment: ERT (imiglucerase, velaglucerase, taliglucerase), SRT (miglustat, eliglustat). Tay-Sachs: HEXA (AR). Hex A deficiency → GM2 ganglioside. Cherry-red spot, neurodegeneration, blindness, seizures, death <4yr. AJ founder. No treatment. Sandhoff: HEXB (AR). Both Hex A+B. Niemann-Pick A/B: SMPD1 (AR). Sphingomyelinase deficiency. Type A: infantile, cherry-red spot (50%), neurodegeneration, death <3yr. Type B: hepatosplenomegaly, lung, no neurologic. Treatment: olipudro alfa (ERT) for B. NPC: NPC1/2 (AR). Cholesterol trafficking. Vertical supranuclear gaze palsy, ataxia, dystonia, dementia, cataplexy, hepatosplenomegaly. Miglustat. Fabry: GLA (X-linked). Alpha-Gal A deficiency → Gb3. Classic male: acroparesthesias, angiokeratomas, cornea verticillata, hypohidrosis, renal failure, HCM, stroke. Females variable. Treatment: ERT (agalsidase alfa/beta), migalastat (chaperone for amenable missense). Hurler (MPS I): IDUA (AR). Alpha-L-iduronidase deficiency. Coarse facies, corneal clouding, dysostosis multiplex, ID, hepatosplenomegaly, cardiac valves, gibbus. ERT (laronidase), HSCT (<2.5yr). Hunter (MPS II): IDS (X-linked). Iduronate sulfatase. No corneal clouding. ERT (idursulfase). Metachromatic leukodystrophy: ARSA (AR). Neurologic, ataxia, spasticity, neuropathy. ERT intrathecal, HSCT. Krabbe: GALC (AR). Infantile: irritability, hypertonia, opsithotonos, decline. HSCT before symptoms.

Zellweger: PEX genes (AR). Peroxisome biogenesis. Hypotonia, seizures, dysmorphic (high forehead), hepatomegaly, renal cysts, chondrodysplasia punctata, elevated VLCFA. Death <1yr. X-ALD: ABCD1 (X). VLCFA accumulation. Childhood cerebral ALD (rapid demyelination, vision loss, behavioral, adrenal insufficiency). Adrenomyeloneuropathy (adult, spastic paraparesis). NBS available. Treatment: HSCT early, gene therapy (eli-cel/Skysona). Lorenzo oil not effective for cerebral. Wilson: ATP7B (AR). Copper accumulation. Hepatic (hepatitis, cirrhosis, fulminant + Coombs-negative hemolysis), neurologic (tremor, dysarthria, dystonia), Kayser-Fleischer rings (copper in Descemet membrane, pathognomonic). Diagnosis: low ceruloplasmin, elevated 24h urinary Cu, K-F rings on slit lamp, liver Cu quantitation. Treatment: chelation (penicillamine, trientine), zinc (maintenance), liver transplant. Alpha-1 antitrypsin deficiency: SERPINA1 (AR). PI*ZZ (Glu342Lys, misfolding, liver accumulation, ~1:2000). Emphysema (panacinar, lower lobe, early onset), cirrhosis, hepatoma, panniculitis. Diagnosis: AAT level, isoelectric focusing (phenotype). Treatment: augmentation therapy (IV AAT), liver transplant, LTOT. Hemochromatosis: HFE (AR). C282Y/C282Y (most common), C282Y/H63D (compound). Iron overload → cirrhosis, DM, bronze skin, hypogonadism, cardiomyopathy, arthritis, HCC. Diagnosis: ferritin, TSAT, HFE genotyping. Treatment: phlebotomy to ferritin 50-100. Cystic fibrosis: CFTR (AR). F508del most common (70% in Europeans). Defective Cl- channel. Meconium ileus, pancreatic insufficiency (fat malabsorption, CF-related DM), recurrent respiratory infections (bronchiectasis, Pseudomonas, ABPA), male infertility (CBAVD), sinusitis, nasal polyps, hepatobiliary disease. Sweat Cl- >60 mmol/L (diagnostic). NBS (IRT/DNA). CFTR modulator therapy: ivacaftor (G551D), tezacaftor/ivacaftor, lumacaftor/ivacaftor (F508del), elexacaftor/tezacaftor/ivacaftor/Trikafta (F508del, most effective, ~90% of CF). Gene therapy in trials. Carrier screening: ACOG pan-ethnic.

DNA methylation: Addition of methyl group to cytosine in CpG dinucleotides. CpG islands (promoters, mostly unmethylated in active genes). CpG shores (regions near islands, tissue-specific methylation). DNMT3A/3B (de novo methylation, establishes patterns in development). DNMT1 (maintenance methylation, copies patterns after replication). TET1/2/3 (demethylation, oxidized 5mC to 5hmC/5fC/5caC). Methylation at promoters → transcriptional silencing (recruit MeCP2, MBD proteins, HDACs). Global hypomethylation in cancer (genome instability). Histone modifications: Histone code: HATs (acetylation, active, H3K9ac, H3K27ac). HDACs (deacetylation, repressive). H3K4me3 (active promoters, Set1/MLL). H3K9me3 (heterochromatin, SUV39H). H3K27me3 (Polycomb repression, EZH2/PRC2). H3K36me3 (elongation, SETD2). H3K79me2 (active, DOT1L). H4K20me3 (heterochromatin). Crosstalk: DNA methylation and H3K9me3 reinforce heterochromatin. PRC2 (H3K27me3) and DNMTs cooperate. Chromatin remodeling complexes: SWI/SNF (BAF, ATP-dependent nucleosome mobilization). Mutations in SMARCB1 (rhabdoid tumors), ARID1A (ovarian clear cell), SMARCA4 (lung). ISWI, CHD, INO80.

Imprinting: ~150 imprinted genes expressed from only one parental allele. ICRs (imprinting control centers) are differentially methylated regions (DMRs). Established in germline, maintained after fertilization, erased in primordial germ cells. Prader-Willi: 15q11-q13 paternal deficiency (SNRPN, NDN, MAGEL2, MKRN3). Maternal UPD 15. Angelman: 15q11-q13 maternal UBE3A loss. Paternal UPD 15. Beckwith-Wiedemann: 11p15.5. ICR1 hypermethylation (IGF2 biallelic, H19 silenced). ICR2 hypomethylation (KCNQ1OT1 active, CDKN1C silenced). Overgrowth, omphalocele, macroglossia, Wilms tumor. Russell-Silver: 11p15.5 ICR1 hypomethylation (IGF2 silenced, H19 active). Maternal UPD 7 (10%). IUGR, triangular face, asymmetry. Transient neonatal diabetes (TNDM): 6q24 paternal UPD. X-inactivation (Lyon hypothesis): Early embryonic random inactivation of one X. XIST lncRNA (X inactive specific transcript) coats X chromosome, recruits PRC2 (H3K27me3). TSIX antisense regulates XIST. Escape from X-inactivation: ~15% of X-linked genes (some biallelic expression, variable). Skewed X-inactivation: non-random distribution. Can occur by chance, selection, or mutation. XIST mechanism: lncRNA, silences X in cis. H3K27me3, macroH2A, DNA methylation, late replication. X-linked disease in females: Skewed inactivation can cause expression (Rett, Fabry, DMD carriers with symptoms).

miRNA biogenesis: RNA Pol II transcribes pri-miRNA (hairpin). Drosha/DGCR8 (Microprocessor) cleaves to pre-miRNA (~70nt). Exportin-5 exports to cytoplasm. Dicer cleaves to ~22nt duplex. RISC loading (Ago2). Guide strand binds target mRNA 3'UTR (seed region, 2-7nt). Translational repression and/or mRNA decay. Functions: Development, differentiation, apoptosis, metabolism, cancer (tumor suppressor miR-15/16, oncomiR miR-21). Therapeutic: Anti-miR (antagomirs), miRNA mimics. siRNA: Exogenous (viral, experimental) or endogenous (from long dsRNA). Perfect complementarity, Ago2 cleaves target mRNA. RNAi pathway. Therapeutic: Patisiran (TTR amyloidosis, siRNA), inclisiran (PCSK9, siRNA). lncRNA: >200nt. XIST (X-inactivation). HOTAIR (HOX, PRC2 recruitment, cancer). MALAT1 (nuclear, splicing, metastasis). NEAT1 (nuclear paraspeckles). H19 (imprinted, oncofetal). FMR4 (CGG repeat lncRNA in Fragile X). piRNA: 24-32nt, Piwi proteins, transposon silencing in germline. Circular RNA (circRNA): Back-splicing, miRNA sponges, regulation. Epitranscriptomics: RNA modifications: N6-methyladenosine (m6A), 5-methylcytosine (m5C), pseudouridine. Writers (METTL3/METTL14), erasers (FTO, ALKBH5), readers (YTHDF1-3). Roles in RNA stability, splicing, translation, cancer.

Rett syndrome: MECP2 (X-linked dominant, male lethal). MeCP2 binds methylated DNA, recruits HDACs, represses transcription. Clinical: normal development 6-18mo, then regression, loss of speech/hand use, stereotypical hand wringing, ataxia, seizures, microcephaly, breathing abnormalities (hyperventilation, apnea). MECP2 duplication syndrome (males): severe ID, hypotonia, spasticity, seizures, recurrent infections. ICF syndrome (Immunodeficiency, Centromeric instability, Facial anomalies): DNMT3B (AR). Hypomethylation of pericentromeric repeats. Combined immunodeficiency, facial dysmorphism, intellectual disability. ATR-X syndrome: ATRX (X-linked). Alpha-thalassemia + mental retardation, X-linked. Facial dysmorphism, microcephaly, genital abnormalities, hemoglobin H inclusions. ATRX protein is chromatin remodeler, deposits H3.3 at telomeres and pericentromeric repeats. Rubinstein-Taybi syndrome: CREBBP (RTS1, ~50%) or EP300 (RTS2, ~3%) (AD). HAT activity. Broad thumbs/great toes, arched eyebrows, low-set ears, beaked nose, severe ID, growth retardation, keloids, increased cancer risk. Sotos syndrome (cerebral gigantism): NSD1 (AD, 90% de novo). Histone methyltransferase. Overgrowth, macrocephaly, advanced bone age, coarse facies (downslanting palpebral fissures, pointed chin, frontal bossing), ID, seizures, scoliosis. Weaver syndrome: EZH2 (AD). Overgrowth, macrocephaly, intellectual disability, camptodactyly. Epigenetic clock: DNA methylation age (Horvath clock, 353 CpGs). Predicts chronological age, age acceleration. Accelerated aging in obesity, HIV, Down syndrome, Werner. Slower in centenarians. Cancer cells: hypomethylated globally + hypermethylated at tumor suppressor promoters.

Developmental origins of health and disease (DOHaD): Early life environment programs adult disease risk via epigenetic changes. Dutch Hunger Winter (1944-45): Prenatal exposure to famine → reduced birth weight, increased obesity, T2D, CVD, schizophrenia in adulthood. Associated with altered DNA methylation at imprinted genes (IGF2) and metabolic genes (POMC, INS, LPL). Maternal diet and supplements: Methyl donors (folate, B12, choline, methionine) influence epigenome. Folic acid reduces NTDs partly via methylation. Endocrine disruptors: BPA, phthalates, DES → altered methylation, metabolic effects, transgenerational inheritance in animal models. Tobacco smoke: In utero → altered methylation at AHRR, CYP1A1, GFI1. Persistent into adulthood. Nutrition and aging: Caloric restriction extends lifespan in many organisms, alters epigenetic patterns. Resveratrol (SIRT1 activator, HDAC). Exercise and epigenetics: Acute exercise alters methylation in muscle (metabolic genes). Regular exercise changes global methylation patterns. Stress and epigenetics: Early life stress → methylation changes in NR3C1 (glucocorticoid receptor), FKBP5, SLC6A4 (serotonin transporter). HPA axis programming. Transgenerational epigenetic inheritance: Epigenetic marks passed to offspring. Controversial in humans. Clear in plants, C. elegans, mice (e.g., agouti viable yellow allele, axin fused). Possible examples: Överkalix (Swedish) studies linking grandparental nutrition to grandchild mortality.

Newborn screening (NBS): RUSP: 35+ core conditions. Dried blood spot 24-48h. MS/MS for metabolic. Pulse ox for critical CHD. Hearing screen. CF, SCD, CH, CAH, PKU, MCAD, GA1, VLCAD. Molecular second-tier for CFTR, ACADM. Carrier screening: ACOG pan-ethnic: CF, SMA, hemoglobinopathies. Ethnic-based: AJ (Tay-Sachs, Canavan, familial dysautonomia, Bloom, Fanconi C, Niemann-Pick A, mucolipidosis IV). Expanded carrier screening (274-600+ genes): higher detection rate, more VUS, counseling implications. Preconception ideal. Cascade screening: When a pathogenic variant identified in proband, offer testing to at-risk relatives (first-degree → second-degree). Critical for Lynch, HBOC, FH, hemochromatosis. Increased detection rate, cost-effective. NIPT in population screening: Cell-free DNA for trisomies 21/18/13. NGS-based. Implementation in public health settings (UK, Belgium, Netherlands). PPV varies: >90% for T21, ~60-80% for T18/13, lower for sex aneuploidies. Polygenic risk scores (PRS): Aggregating thousands of small-effect variants. Clinical utility emerging: breast cancer (combined with mammography risk stratification), CAD (reclassify intermediate risk), T2D. Implementation challenges: population transferability (Euro-centric bias), clinical actionability, education of providers/patients, regulation. Precision medicine and EHR: Integration of genomic data (PGx, carrier, diagnostic) into electronic health records. Clinical decision support (CDS) alerts for PGx (CPIC guidelines). Pre-emptive PGx panel (e.g., rightPGx, PG4KDS) before prescribing. CPIC guidelines: 100+ gene-drug pairs with dosing recommendations. Level A (CPIC, DPWG): strong recommendation. Implementation in health systems: pilot studies, pharmacist-led, EHR alerts.

ctDNA: Cell-free tumor DNA in blood. Shed from apoptotic/necrotic tumor cells. Half-life ~30-120min. Detection methods: digital PCR (ddPCR, BEAMing for known mutations), NGS (targeted panels, WES, WGS for comprehensive profiling). Applications: Molecular profiling when tissue inadequate (Guardant360, FoundationOne Liquid). MRD detection (signatera, bespoke panels). Treatment response monitoring (ctDNA levels correlate with tumor burden). Resistance mutation detection (EGFR T790M, KRAS G12C resistance). Early cancer detection (multi-cancer early detection MCED: Galleri, CancerSEEK). Multi-cancer early detection: Methylation-based (cfDNA fragmentomics, methylation patterns tissue-specific). Galleri (targeted methylation, ~50 cancers, tissue of origin ~90% accuracy). Sensitivity stage-dependent (I ~20-40%, IV ~90%). Specificity >99%. Implementation considerations: false positives, overdiagnosis, cost, follow-up protocols. MRD in solid tumors: ctDNA after curative intent treatment. Positive MRD predicts recurrence (months to years before imaging). Lead time: 3-12mo for most cancers. Clinical trials: ctDNA-guided adjuvant therapy (DYNAMIC, CIRCULATE). Liquid biopsy challenges: Low ctDNA fraction (early stage, brain tumors), clonal hematopoiesis of indeterminate potential (CHIP), standardization, reimbursement.

Gene therapy approaches: Gene addition (AAV, lentivirus), gene editing (CRISPR, base editing, prime editing), gene silencing (ASO, siRNA), gene replacement (large genes, dual AAV). AAV vectors: AAV2, AAV8, AAV9. Serotypes determine tropism. Capsid engineering. Low immunogenicity (but pre-existing antibodies). Non-integrating (episomal). Limitations: packaging capacity ~4.7kb, dilution in dividing cells. Approved AAV therapies: Luxturna (AAV2, RPE65, inherited retinal dystrophy). Zolgensma (AAV9, SMN1, SMA <2yr, single IV dose). Elevidys (AAV, DMD, microdystrophin). Hemgenix (AAV5, FIX, hemophilia B). Roctavian (AAV5, FVIII, hemophilia A). Lentiviral vectors: Integrating (risk of insertional mutagenesis). Ex vivo: CAR-T cells (tisagenlecleucel, axicabtagene ciloleucel, brexucabtagene, lisocabtagene). Anti-CD19 for B-ALL/DLBCL, BCMA for multiple myeloma. Gene corrected HSCs: betibeglogene autotemcel (Zynteglo, beta-thalassemia), exagamglogene autotemcel (Casgevy/exa-cel, sickle cell). CRISPR therapies: Casgevy (exa-cel): ex vivo CRISPR-Cas9 edited CD34+ HSCs (BCL11A enhancer, reactivates fetal Hb). Approved for sickle cell disease and beta-thalassemia. In vivo editing: LCA10 (EDIT-101, CRISPR, CEP290, trial). Transthyretin amyloidosis (NTLA-2001, in vivo CRISPR with LNP, liver editing). Base editing: Adenine base editor (ABE, A>G). Cytosine base editor (CBE, C>T). No DSB needed. BEAM-101 for sickle cell. Prime editing: Cas9 nickase + reverse transcriptase + pegRNA. Precise edits (all 12 possible base substitutions, small indels). Larger edits possible (twin prime). ASO/siRNA therapies: Nusinersen (Spinraza, SMA, ASO). Tofersen (QALSODY, SOD1 ALS). Patisiran (Onpattro, TTR amyloidosis, siRNA). Inclisiran (Leqvio, PCSK9, siRNA). Germline editing: Controversial and ethically fraught. International moratorium. 2018 He Jiankui case (CCR5-edited twins Lulu and Nana) condemned. Somatic vs germline: Somatic: acceptable (consent required, non-heritable). Germline: not currently acceptable (off-target, ethics, heritable changes). Genetic counselling: Non-directive, respect patient autonomy, informed consent, psychological support, family communication, cascade testing, reproductive options (prenatal, PGD, adoption, donor gametes). GINA (2008): prohibits genetic discrimination in health insurance and employment (does not cover life/disability/LTC insurance). State-level variations. Ethical issues: VUS, incidental findings, recontact, privacy, data sharing, equity, justice, access, cost. Duty to warn relatives varies by jurisdiction.