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Introduction to Cytogenetics
Readings
Jorde, Carey, Bamshad & White: Medical Genetics, 3rd edition, C.V. Mosby Publishing, 2005.
- Cell Cycle pp. 22-27
- Chapter 6 p. 107
Objectives
- Normal karyotype: number of chromosomes, basic nomenclature
- Typical indications for chromosome analysis in the newborn and adult. Examples of the findings
- Meiosis and the errors that can lead to trisomy/mis-segregation of balanced translocations
How to See Chromosomes
Chromosomes only visible as cells approach and pass through metaphase in cell cycle
What is seen is DUPLICATED chromosome
Normal human karyotype has 46 chromosomes - 22 pairs of autosomes and a pair of sex chromosomes
- Female 46,XX
- Male 46,XY
Karyotype Arrangement
Chromosomes have characteristic banding patterns created by trypsin and Giemsa
Chromosomes laid out in pairs, from largest (#1) to smallest (#21)
Arranged with short arm (p from French “petit”) going up, and long arm (q) going down
Chromosome movement via centromere
Nomenclature
International System for Human Cytogenetic Nomenclature (ISCN)
Karyotype components:
- Number of chromosomes, sex, first change, second change, etc.
- Sex chromosome changes are listed first and then others in numerical order
Monosomy
Loss of one copy of a chromosome
45,X known
No other full monosomies reported
Loss of part of a chromosome = monosomy for small region
Example 1:
Trisomy
Three copies of a chromosome or part of a chromosome - normal chromosome number increased by 1 to 47
Triploidy
Three copies of EVERY chromosome - normal chromosome number increased by 23 to 69
Typical Newborn Presentation
Baby G is admitted to Neonatal Intensive Care Unit. Staff notes that baby has unusual features and call for Genetics consult. Genetics find the baby has: Hypotonia, protruding tongue, slanted palpebral fissures, small nose with low nasal bridge, small ears, short neck, relatively short metacarpals and phalanges in hands, wide gap between first and second toes (“sandal gap”)
In the Lab
0.25 – 0.5 ml blood added to 5 ml tissue culture medium (with fetal calf serum) Phytohemagglutinin (PHA) added to stimulate T-cells to divide. First harvest done after 48 hours
Back to the Blood in Culture - Harvest:
- Colchicine is added to disrupt mitotic spindle
- Hypotonic solution added to swell the cells
- Cells fixed
- Cell suspension dropped onto slides
- Slides are treated with trypsin solution and stained (G-banding)
How Does Trisomy 21 Happen?
- Gametes form during meiosis
- In meiosis 1 (MI) homologous chromosomes go to different daughter cells
- In meiosis 2 (MII) each chromosome divides along its length and each chromatid goes to a daughter cell
Frequency of Trisomy
| Trisomy | Frequency | Parental Origin |
| 47,Xx/Xy,+21 | 1 In 700 | Mat MI>Mat MII 3:1 |
| 47,Xx/Xy,+18 | 1 In 6 – 8,000 | Mat MI Or MII |
| 47,Xx/Xy,+13 | 1 In 12,000 | Mat MI Or MII |
Typical Routine Adult Presentation
Mr. and Mrs. F, who are in their late 20’s, are referred to a fertility clinic because they have been married for 5 years, but have no children. Mrs. F has been pregnant four times, but all pregnancies have ended in miscarriage. As part of their workup, blood is sent for karyotype analysis. In the lab blood is cultured as before, but for 72 hours. Mr. F is found to have normal karyotype, 46,XY. Mrs. F is found to have apparently balanced translocation between chromosomes 10 and 11.
Mrs. F’s Karyotype 46,XX,t(10;11)(p15;q21)
- Translocations = exchange of material between two or more chromosomes
- Constitutional balanced translocation usually has no effect on phenotype
- May result in infertility or history of spontaneous abortion
- Constitutional translocations are present at birth in some or all of cells in body and most are unique to a family
- Robertsonian translocations occur between acrocentric chromosomes (13/14/15/21/22) Results in loss of non-critical genes in the short arm regions of the chromosomes involved
Robertsonian translocation
Count is decreased to 45: 45,XY,der(15;22)(q10;q10) or 45,XX,der(13;14)(q10;q10)
“q10” means that the break is in the centromere and the long arm is present
Frequency of Robertsonian Translocation Combinations
| 13 | 14 | 15 | 21 | 22 | |
| 13 | X | 45% | 3% | 6% | <1% |
| 14 | X | 1% | 36% | <1% | |
| 15 | X | 3% | 1% | ||
| 21 | X | 4% |
Translocations and Meiosis
- Chromosomes pair along their lengths as far as possible making a four - armed figure
- Centromeres of homologs repel each other and go to different daughter cells
- Segregation errors may lead to combination of monosomy and trisomy
Mr. and Mrs. F decide to try to get pregnant one more time. Pregnancy ends in spontaneous loss, and tissue sent to the lab.
Karyotype of Fetus
46,XX,der(10)t(10;11)(p15;q21)
- Unbalanced translocations will impact phenotype due to a mixture of monosomy and trisomy for the regions involved
- Unbalanced translocations may be incompatible with life
- Written as: 46,XX,der(5)t(5;8)(p13;q22)
Summary
- Normal karyotype = 46 chromosomes. 22 pairs of chromosomes and 2 sex chromosomes (46,XX or 46,XY)
- Chromosomes divided by the centromere into short (p) and long (q) arms
- Each chromosome has a number of bands, each of which has been assigned a number
- Karyotypes written using a system of nomenclature
- Abbreviations used to describe changes such as t, der, dup, inv
- Balanced constitutional rearrangements (translocation or inversion) do not alter phenotype and most are unique to a family
- Unbalanced constitutional rearrangements result in duplication and deletion of material and are associated with changes in phenotype
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