1. Dr. Nasser A Elhawary Professor of Medical Genetics
Chromosomal Abnormalities
Dr. Nasser A Elhawary
Professor of Medical Genetics

2. 2 Chromatides Centromere
A chromosome distincts into 2 “sister chromatides”.
They are linked together via a centromere

3. Tjio & Levan reported it is 46 chrom. in 1956
Human Chromosomes…
Contains DNA and Protein
46 chromosomes
Autosomes: Pairs 1-22
Sex chromosmes
Human metaphase

4. Identification of Chromosomes…
The chr’s are present in pairs ‘homologs’ (one from female, the other from male).
Diploid cells (2n): Cells that contain pairs of homologous chr’s.
Haploid cells (n): Certain cells (sperm/egg gametes) contain only one copy of each chr.
At fertilization, the fusion of haploid gametes together produces a cell carrying the diploid no. of chr’s (zygote).

5. Classification of Chromosomes
The centromere (1ry constriction) divides the chr into 2 arms (petit short ‘p’-arm & long ‘q’-arm).
The location of centromere in each chr is characteristic for a given chromosome.
‘Metacentric’: a chromosome with a centrally placed centromere.
‘Submetacentric’: a chromosome with a centromere close to one end than the other.
‘Acrocentric’: a chr with the centromer placed very close to one end.

6. Classification of Chromosomes…

7. Chromosome Preparation

8. Human chromosomes are arranged in groups from A to G.
Chromosome Banding…
Human chromosomes are arranged in groups from A to G.
Each group is defined by chromosomal size and centromere location.
In 1960s, new staining procedures were developed that resulted in banded chromosomes (‘dark’ and ‘light’).
Banding protocols: G-banding, R-banding, Q-banding, C-banding, etc.

9. G-banded Human Chromosomes
Chromosome Banding… A
karyptype of
G-banded Human Chromosomes

10. R-banding…

11. Q-banding

12. C-banding…

13. What is the importance of high resolution banding?

14. Identification of Chromosomes…
Each arm is subdivided into numbered regions starting at the centromere.
Thus, any region can be identified by a descriptive address such as Xq27.3/
FMR1 gene

15. Karyotype Analysis…
The banding pattern of the chromosomes in the human karyotype.

16. Banding chromosomes…
G-Banding using trypsin and Giemsa stain the chromatin in 2 main forms:
Euchromatin: stains ‘light’ and consists of genes which are actively expressed.
Heterochromatin stains ‘dark’ and is made up largely of inactive unexpressed repetitive DNA.

17. Molecular Cytogenetics
Fluorescence In-Situ Hybridization (FISH):
It is ability of a portion of ssDNA (i.e. probe) to anneal with its complement-ary target sequence on: i) a metaphase chromosome, ii) interphase nucleus or iii) extended chromatin fiber.

18. FISH is widely used in clinical diagnostic purposes.
In FISH, the DNA probe is labeled with a fluorochrome + patient’s chrom visualized using fluorescent microscope.

19. Types of FISH 1- Centromeric probes.
2- Chromosome-specific unique-sequence probes.
3- Telomeric probes.
4- Whole chromosome paint probes.

20. Types of FISH… 1- Centromeric probes
FISH of interphase nuclei with Centromeric probes for chromosomes 18.X and Y: showing 3 signals consistent with trisomy 18.

21. 2- Unique sequence probes are useful for identifying tiny submicroscopic deletions & duplications.
Also, use of interphase FISH probe to identify HER2 over-expression in breast tumors.
Metaphase image: Chromosome band 7q11.23 showed deletion associated with Williams syndrome. Normal chrom. has 2 signals (green) for the control probe and the ELN gene probe signal (orange), but the deleted chrom. shows only the control probe signal

22. Types of FISH… Telomeric probes
Telomeric probes have been used for identifying tiny ‘cryptic’ subtelomeric abnormalities, e.g. deletions, translocations.

23. Types of FISH… 3- Whole-chrom paint probes
These consists of a cocktail of probes obtained from different parts of a particular chromosome.
When this mixture of probes is used together in a single hybridization, the entire chromosome fluoresces (i.e. is “painted”).
Useful in rearrangements (e.g. translocations) & additional chromosomal materials.

24. Types of FISH… Whole-chrom paint probes
Chrom painting showing a reciprocal translocation involving chrom 3 (red) & 20 (green).

25. Types of FISH… Whole-chrom paint probes
M-FISH or Spectral karyotyping (SKY) uses pools of whole human chrom. paint probes to provide a multicolor human karyotype. Each homologous chrom. shows a unique color.
These are useful to detect chromosomal rearrangements (e.g. deletions, trans-locations), ring chrom, …

26. Types of FISH… Whole-chrom paint probes
M-FISH showing complex chromosome rearrangement involving chromosomes 4, 8, 13, 18, and 21

27. Genomic Hybridization
Comparative
Genomic Hybridization
CGH was originally developed to overcome the difficulty of obtaining good-quality metaphase preparations from solid tumors.
CGH enables the detection of regions of allele loss and gene amplification.
Tumor ‘test’ DNA is labeled with green paint & control normal DNA with red paint.
The two samples are mixed and hybridized com-petitively to normal metaphase chromosomes.
watch the green-to-red ratio????

28. Genomic Hybridization
Comparative
Genomic Hybridization
CGH Analysis showing areas of gene amplification & reduction (del) in tumor DNA.
DAPI: diamidophenylindole
FITC: Fluorescein isothiocyanate

29. CGH limits for more than 10Mb for losses and 2Mb for gains.
CGH extended to include the analysis of single cells for prenatal diagnosis following whole-genome amplification.
CGH limits for more than 10Mb for losses and 2Mb for gains.
Microarray or array CGH is likely to replace metaphase CGH.

30. Chromosomal Abnormalities
Numerical
Structural

31. Numerical Abnormalities
Aneuploidy: Loss or gain one or more chromosome
Polyploidy: Addition of one or more complete haploid sets (69, 92 chromosomes).
Monosomy: Loss of a single chromosome (Turner syndrome: 45,X0)
Trisomy: gain of a single chromosome (e.g. Down syndrome: 47,XY,+21).

32. Aneuploidy.. Numerical abnormality…
Turner syndrome
Monosomy
Down syndrome
Trisomy 21

33. A Karyotype of triploidy cell, 69,XXX

34. Mechanism of Trisomy

35. Structural Abnormalities
Translocation: Transfer a genetic material from one chromosome to another.
Reciprocal translocation: Breakage in 2 chromosomes and exchanged.
Robertsonian translocation: Breakage close to centromeres of acrocentric chrom. (e.g. Down syndrome)

36. Translocation

37. Origin of translocation
in mother giving Down syndrome

38. Structural abnormalities…
4- Deletions: loss of part of a chromosome
(e.g. Wolf syndrome ‘4p-’, cri du chat syndrome ‘5p-’).
5- Insertions: a segment of one chromosome is inserted into another chromosome.
6- Inversions: a two-break rearrangement in a chrom. which re-inserted in the inverted positions.
7- Ring chromosome: 2 breaks leaving sticky ends which re-unit to give ring chromosome.
8- Isochromosomes: from loss of one arm of the chrom. with duplication of the other.

39. Cri du chat syndrome (5p–)

40. Insertion

41. Ring chromosome
(1926)

42. Inversion:
A) Pericentric inversion
B) Paracentric inversion

43. Mosaicism & Chimerism
Mosaic: Presence of two or more cell lines in an individual, but derived from the same zygote.
Mosaic produces from non-disjunction in an early embryonic mitotic division (mosaic trisomy 21).
Chimerism: Presence of two or more cell lines in an individual, derived from more than one zygote.
Blood Chimeras & Dispermic chimeras.

44. Mosaicism & Chimersim mechanism