Molecular Techniques - Chromosomes

Question 1

What is karyotyping?

Answer 1

i) Prepare metaphase cells and stain the DNA to highlight DNA banding patterns.
ii) Karyotype to line up according to chromosome number.

Question 2

What is FISH?

Answer 2

• Fluorescent In Situ Hybridisation

- Make specific fluorescently-labelled DNA probes and expose to chromosome within the cell.

Question 3

What does FISH allow the investigation of?

Answer 3

Macro mutations, e.g. Deletions, insertions, translocations, etc.

Question 4

What is chromosome painting?

Answer 4

• Make a families of probes of different colours, each specific to each chromosome.
• Can be used to compare healthy and cancer genomes, e.g. Many translocations in cancer cells.

Question 5

What is cytogenetics?

Answer 5

Study of genetic constitution of cells through the visualisation and analysis of chromosomes.

Question 6

Why is one referred to a cytogeneticist?

Answer 6

1. Acquired abnormalities (e.g. Leukaemias, solid tumours)
2. Constitutional abnormalities
   - prenatal diagnosis
   - birth defects
   - abnormal sexual development
   - infertility
   - recurrent foetal loss

Question 7

What are the 3 main types of prenatal diagnosis methods?

Answer 7

1. Ultrasound scans
2. Maternal serum screening (looks for biochemical markers)
3. DNA/chromosome analysis

Question 8

Compare the 2 methods used for obtaining pre-natal DNA samples.

Answer 8

1. Chorionic villus sampling
   - 11-12 wks gestation
   - 1.2% miscarriage risk
   - Sample of chorionic villi removed from placenta
2. Amniocentesis
   - >15 wks
   - 0.8% miscarriage risk
   - Sample of amniotic fluid removed from sac

Question 9

How are karyotype reports formatted?

Answer 9

• Chromosome number, sex complement, structural changes
• E.g. 46,XX
  47,XY,+21

Question 10

What can karyotyping be used to detect?

Answer 10

1. Aneuploidy/polyploidy
2. Rearranged chromosomes (only see differences in size)
3. Chromosome activity (euchromatin stains differently to heterochromatin)

Question 11

What can FISH be used to detect?

Answer 11

1. Aneuploidy (centromere probes)
2. Microdeletions/duplications (locus/gene specific probes, telomere probes)
3. Rearranged chromosomes (whole chromosome paints)

Question 12

Why is FISH often better to use for PND than karyotyping?

Answer 12

• Karytotyping can take up to 14 days in culture (causes anxiety).
• FISH probes for 13, 18, 21, X and Y using uncultured cells allows results in 24-48 hrs. Many patients TOP after FISH.

Question 13

What is microarray comparative genomic hybridisation (aCGH) used to detect?

Answer 13

• Aneuploidy

- Unbalanced karyotypes - but does not detect balanced rearrangements

Question 14

aCGH is used for the diagnosis of patients with what kind of symptoms?

Answer 14

Learning difficulties, developmental delay, multiple congenital abnormalities…