What Is a Chromosome?A long, continuous strand of DNA, plus several

types of associated proteins, and RNA.

• Cytogenetics: the study of chromosomes, their structure, and their inheritance.

• The importance of chromosomes in medicine:– Clinical diagnosis

• Eg, Turner syndrome

– Gene mapping• Eg, cystic fibrosis

– Cancer cytogenetics• Eg, retinoblastoma

– Prenatal diagnosis• Eg, Down syndrome

Stalkqp

21

17

ACROCENTRICMETACENTRIC SUBMETACENTRIC

Stalk

CentromereSatellite

qLongArm(q)

ShortArm(p)

pp

q

213

– Size:

» chromosome 1 is largest, chromosome 22 smallest

– Centromere position:

» metacentric, submetacentric, acrocentric

» all chromosomes have a shorter arm, p, and a longer arm, q.

Karyotype: a picture of all the chromosomes arranged by type:“Karyotype” also refers to the standard chromosome set of an individual or a species.

• G-banding: – pattern subdivides

each chromosome arm into regions.

1. Cells (from blood, amniotic fluid, or chorionic villus) are grown in culture.Mitogens may be required: lymphocytes require phytohemagglutinin

2. Colcemid stops cells at metaphase.

3. Hypotonic shock ruptures RBCs, swells lymphocytes.

4. Cells are fixed in MeOH/HOAc.

5. Chromosomes are spread on a slide.

6. Trypsinization and staining with Giemsa reveals G-bands.

7. The chromosome spread is photographed and arranged by type.

Karyotypes are made from metaphase chromosomes:

Ideogram of G-banding pattern at 450-band stage:

High-resolution bandings of 550-850 bands can be made from prometaphase chromosomes.

Introduction of molecular cytogenetics:

• DNA in chromosome spreads is denatured and hybridized to probes:– FISH: fluorescent in situ hybridization

• Probes for specific genes or chromosomal regions– Rearrangements, deletions, abnormal chromosome number

– Chromosome painting• Mixture of fluorescently-labeled probes for single-copy genes that

map along the entire length allow entire chromosomes to be visualized

– Anueploidies, translocations

– SKY: spectral karyotyping• All chromosomes painted at once, each chromosome a different color

FISH : gene-specific probes

• Pinkel D, Straume T, Gray JW. Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization. Proc Natl Acad Sci U S A. 1986;83:2934-2938.

•A, Partial metaphase showing a microdeletion of the elastin gene (ELN) (arrow) associated with Williams syndrome.

•B, Interphase cell shows a microduplication of the peripheral myelin protein 22 gene (PMP22) (arrows) associated with Charcot-Marie-Tooth syndrome.

SKY: chromosome-specific probes

Normal karyotype Cancer cell karyotype note: translocations, polysomy, monosomy

Cells used for karyotyping:• Blood cells:

– T-lymphocytes from peripheral blood are easily collected• Growth in culture requires mitogens (phytohemagglutinin)• Cultures are short-lived

• Skin cells– Fibroblast cultures grow without mitogens

• Cultures are long-lived, can be immortalized

• Tumor cells– Obtained from biopsy

• Amniocytes– From amniocentesis

• Amniotic fluid is available after ~16 weeks• Amniotic fluid is waste containing cells shed from skin, respiratory tract, urinary tract.• Amniotic fluid can be biochemically tested for metabolic disorders

• Placental cells– Obtained by chorionic villus sampling

• Placental cells are dividing, and karyotypes can be obtained within hours

Indications for chromosome analysis:

– Family history can indicate a need for prenatal testing:

Amniocentesis and chorionic villus sampling pose small risks of infection or inducing spontaneous abortion, so samples are taken only if indicated by a risk factor.

• Known chromosomal abnormality of first-degree relative

• Previous child with chromosomal aberration– Recurrence risk ~1-2%

• Advanced maternal age: >35 years

• Mother carries X-linked disorder– Male offspring would be at risk

• Prenatal testing:

Chorion

Bladder

Catheter

Uterus

Developingplacenta

Rectum

Amnioticcavity

Chorionicvilli Ultrasound to monitor

procedure

Developing fetus

Amniocentesis: Chorionic villus sampling:

• Other reasons for chromosome analysis:– Fertility problems: infertility or repeated miscarriage

• Chromosomal abnormalities seen in one or both parents at 3-6%– Translocation or inversion

– Stillbirth or neonatal death• Chromosomal abnormalities at ~10%

• Karyotyping important for generating family history

– Problems with a child• Failure to thrive, developmental delay, multiple malformations,

ambiguous genitalia, mental retardation

– Neoplasia: all cancers show chromosomal abnormalities• Karyotype can provide diagnostic or prognostic information

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