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Genetics Charts - Answers

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SLIDE 1

1. The subject rinses the mouth thoroughly with water to remove food debris.
2. A clean wooden spatula or cytobrush is used to gently scrape the inner surface of the cheek (buccal mucosa).
3. The collected material is immediately smeared onto a clean glass slide.
4. The smear is fixed immediately in equal parts of 95% ethanol and ether (or simply 95% ethanol) for 30 minutes to prevent drying artefacts.
5. The fixed smear is stained with cresyl violet or crystal violet (or haematoxylin) to demonstrate the Barr body.
6. The slide is examined under oil immersion. The Barr body appears as a darkly staining plano-convex mass, 1 µm in diameter, applied to the inner surface of the nuclear membrane.
7. At least 100 well-preserved nuclei are examined; in normal females, ≥20-30% of nuclei show a Barr body.

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1. One X chromosome is genetically active in each somatic cell of a female.
2. The other X chromosome (either maternal or paternal origin) undergoes heterochromatization and is rendered genetically inactive - forming the Barr body.
3. Inactivation is random - in each cell of the blastocyst (~day 5.5 of embryonic life / ~16-cell stage), either the maternal or the paternal X is inactivated randomly.
4. Inactivation is permanent and heritable - the same X chromosome remains inactivated in all daughter cells derived from that precursor cell.
5. As a result, normal females are mosaics - one population of cells has the maternal X inactivated, the other has the paternal X inactivated.

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SLIDE 2

• The proband/propositus is the first affected family member who brings the family to medical attention; indicated by an arrow.
• Pedigree charts are used to determine the pattern of inheritance (autosomal dominant, autosomal recessive, X-linked, etc.) and to assess recurrence risk.
• Generations are labeled with Roman numerals (I, II, III...) and individuals within a generation with Arabic numerals.

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SLIDES 3-8: Karyotypes

1. Dividing cells (lymphocytes from peripheral blood, amniotic fluid cells, or chorionic villi) are cultured.
2. Cell division is arrested at metaphase using mitotic spindle inhibitors (e.g., colchicine/N-diacetyl-N-methylcolchicine).
3. Cells are treated with a hypotonic solution to swell them and spread chromosomes.
4. Fixed and stained - most commonly with Giemsa stain (G-banding), producing characteristic alternating light and dark bands on each chromosome.
5. Chromosomes are arranged in pairs according to size (largest to smallest), centromere position, and banding pattern.
6. A karyotype is expressed as: [total chromosome number], [sex chromosome complement], [any abnormality]
7. Standard G-banding resolves 400-800 bands per haploid set; prophase banding up to 1500 bands.

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• Monosomy - loss of one chromosome (2n-1), e.g., 45,X (Turner)
• Trisomy - extra chromosome (2n+1), e.g., trisomy 21 (Down), trisomy 18 (Edwards), trisomy 13 (Patau)
• Polyploidy - complete extra haploid set (triploidy = 3n=69, tetraploidy = 4n=92)
• Mosaicism - two or more genetically distinct cell populations

• Deletions, duplications, inversions, translocations (reciprocal/Robertsonian), isochromosomes, ring chromosomes

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SLIDE 4 - Karyotype Identification

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SLIDE 5 - Karyotype Identification

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SLIDES 6-8 - Karyotype Identification

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SLIDE 9

• Satellite chromosomes: Acrocentric chromosomes (13, 14, 15, 21, 22) have small chromatin masses (satellites) attached to the short arm by a secondary constriction (stalk) - these contain rRNA genes
• Size: The 23 pairs are numbered 1-22 from largest to smallest, plus sex chromosomes
• Denver system (1960): Groups A (1-3), B (4-5), C (6-12,X), D (13-15), E (16-18), F (19-20), G (21-22,Y)

• p arm = short arm (from French "petit")
• q arm = long arm
• Centromere = primary constriction; point of spindle fiber attachment
• Telomere = chromosome tip; protective cap

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SLIDES 10-13: Syndrome Identification

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SLIDE 10

• Round, flat face with upward-slanting palpebral fissures (mongoloid slant)
• Epicanthic folds (Brushfield spots on iris)
• Flat nasal bridge
• Small ears, protruding tongue (macroglossia relative to small mouth)
• Single palmar crease (simian crease)
• Brachycephaly (flat occiput)

• Short stature
• Short broad neck
• Hypotonia (floppy infant)
• Short stubby fingers; clinodactyly (incurved 5th finger)
• Wide gap between 1st and 2nd toes ("sandal gap")

• Cardiac: Congenital heart defects in 40-50% (VSD, ASD, endocardial cushion defect/AVSD)
• GI: Duodenal atresia ("double bubble" sign), Hirschsprung disease
• Hematology: 10-15x increased risk of leukemia (especially AML-M7 and ALL)
• CNS: Intellectual disability (IQ 25-50), early-onset Alzheimer disease (virtually all >40 years develop AD neuropathology)
• Endocrine: Hypothyroidism, increased risk of diabetes
• Immune: Increased susceptibility to infections

• Most common (95%): Trisomy 21 - 47,XX,+21 or 47,XY,+21 (nondisjunction during meiosis I, maternal origin in ~95% of cases; risk increases with maternal age)
• Translocation (4-5%): 46 chromosomes but extra chromosome 21 material attached to another chromosome (usually chromosome 14) - Robertsonian translocation; 46,XX,rob(14;21)(q10;q10),+21 - does NOT increase with maternal age; recurrence risk is high
• Mosaicism (1-2%): 46/47,+21 - milder phenotype

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SLIDE 11

• Eunuchoid body habitus - long limbs, span > height, increased lower segment
• Small, firm, atrophic testes (most consistent finding; typically < 3.5 cm in adults)
• Azoospermia and infertility (most common cause of male infertility due to a chromosomal disorder)
• Gynecomastia (in ~50%)
• Reduced body/facial hair
• Small penis
• Tall stature with disproportionately long legs
• Reduced muscle mass
• Subtle cognitive deficits - particularly verbal/language skills
• Increased risk: type 2 diabetes, metabolic syndrome, mitral valve prolapse (50%), osteoporosis, breast cancer (20x increased risk vs. normal males), extragonadal germ cell tumors
• Elevated FSH and LH; low testosterone

• Classic: 47,XXY (90% of cases) - extra X due to nondisjunction in either parent during meiosis
• Variants: 48,XXXY; 49,XXXXY; 48,XXYY; 46,XY/47,XXY (mosaic)
• The greater the number of supernumerary X chromosomes, the more severe the phenotype and intellectual disability

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SLIDE 12

• Lymphedema of dorsum of hands and feet at birth
• Cystic hygroma (neck swelling due to distended lymphatics)
• Webbing of the neck (pterygium colli) - from resolved cystic hygroma
• Short stature (rarely exceeds 150 cm) - most common presenting feature
• Shield chest with widely spaced nipples
• Broad, "shield-shaped" chest

• Congenital heart disease in 25-50%
• Coarctation of aorta (most characteristic - 15-20%)
• Bicuspid aortic valve (most common cardiac defect - 30%)
• Aortic root dilatation (30%); 100-fold increased risk of aortic dissection

• Primary amenorrhea (most common cause - accounts for 1/3 of all cases)
• Failure of secondary sex characteristics at puberty
• Streak gonads (fibrotic ovaries) - ovarian dysgenesis
• Infertility
• Infantile genitalia; minimal pubic hair; poor breast development

• Cubitus valgus (increased carrying angle)
• Low posterior hairline
• Pigmented nevi
• Renal anomalies in 30-50% (horseshoe kidney most common)
• Normal intelligence (subtle deficits in visual-spatial processing)
• Sensorineural hearing loss

• 57% : 45,X (monosomy X) - most common and most severe; results from nondisjunction (paternal gamete usually lacks sex chromosome in ~70% of 45,X cases)
• Structural abnormalities (14%): Isochromosome X long arm [46,X,i(Xq)]; ring chromosome [46,X,r(X)]; deletions [46,X,del(Xp) or del(Xq)]
• Mosaicism (29%): 45,X/46,XX (milder phenotype); 45,X/46,XY (risk of gonadoblastoma - gonadectomy advised); 45,X/47,XXX

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SLIDE 13

• Clenched fists with overlapping fingers (index finger over 3rd, 5th finger over 4th) - pathognomonic
• Rocker-bottom feet (prominent calcaneus)
• Low birth weight; severe growth retardation
• Microcephaly with prominent occiput
• Small mouth, micrognathia
• Low-set, malformed ears
• Cardiac defects (95%): VSD, ASD, PDA
• Renal malformations
• Diaphragmatic hernia
• Omphalocele
• Severe intellectual disability
• Survival: median ~10 days; <10% survive to 1 year

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• Holoprosencephaly (failure of forebrain to divide)
• Microphthalmia / Anophthalmia
• Cyclopia (in severe cases)
• Cleft lip and/or palate
• Polydactyly (post-axial)
• Microcephaly
• Cardiac defects (80%): VSD, ASD, PDA, dextrocardia
• Scalp defects (cutis aplasia)
• Renal cysts
• Capillary hemangiomas
• Severe intellectual disability
• Survival: median 7-10 days; <5% survive to 6 months

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Summary Table: Key Chromosomal Syndromes