Clinical Genetics: Chromosomal abnormalities I Flashcards
1
Q
What form of DNA do chromosomes usually exist as?
A
- Chromosomes usually exists as chromatin
- DNA double helix wound around an octamer of histone proteins
- Octamer of histones form nucleosome
- Nucleosomes packaged together with scaffolding proteins to form chromatin
2
Q
What is the difference between euchromatin and heterochromatin?
A
-
Euchromatin
- Uncondensed, dispersed through nucleus
- Allows gene expression
-
Heterochromatin
- Highly condensed, genes not expressed
3
Q
DNA is usually loosely packaged within the chromosome. When is this not the case?
A
- Not the case during cell division when DNA is complexed with various proteins and undergoes several levels of compaction through coiling and supercoiling
4
Q
What are homologous chromosomes?
A
- Homologous chromosomes are a pair of identical chromosomes, same length, genes and centromere position.
- One of the pair of chromosomes is inherited from your mother and the other inherited from your father
5
Q
What is a gene locus?
A
- A gene locus is the location of a particular gene on a chromosome
6
Q
What is an allele?
A
- An allele is an alternate form of a gene
- At each gene locus an individual has 2 alleles, one from each homologous chromosome
7
Q
Why are chromosomes sometimes shown with a single chromatid?
A
- Chromosomes with single chromatid show how chromosomes look during interphase - after cell division
8
Q
Why are chromosomes sometimes shown with two sister chromatids?
A
- Chromosomes with 2 sister chromatids show how chromosomes look after S phase where DNA is duplicated in anticipation of cell division
9
Q
Briefly describe the stages of the cell cycle
A
- G1 - Cellular contents, except chromosomes are duplicated, Cell makes proteins needed for DNA replication
- S phase - Chromosomes are replicated so that each chromosome now consists of two sister, identical chromatids
- G2 - Synthesis of proteins especially microtubules
- Mitosis - Cell divison
10
Q
How many pairs of chromosomes do humnas have?
A
- 23 pairs of chromosomes
- 22 pairs autosomes, 1 pair sex chromosomes XX or XY
11
Q
What are the 3 different types of chromosome? State which chromosomes within the human genome belong to each type of chromosome
A
-
Metacentric - p & q arms even length
- 1-3, 16-18
-
Submetacentric - p arm shorter than q
- 4-12, 19-20, X
-
Acrocentric - Long q, small p; p contains no unique DNA
- 13-15, 21-22, Y
12
Q
What are the different types of chromosomal changes and how can each type be detected?
A
-
Numerical changes - Can be detected through:
- Traditional karyotyping
- FISH
- QF-PCR (Quantitative fluoresence PCR)
- NGS
-
Structural changes - Can be detected through:
- Traditional karyotyping
- FISH
13
Q
What is meant by the term “Haploid”?
A
- One set of chromosomes (n=23) as in a normal gamete
14
Q
What is meant by the term “Diploid”?
A
- Cell contains two sets of chromosomes (2n=46; normal in human)
15
Q
What is meant by the term “Polyploid”?
A
- Any chromosome number which is an exact multiple of the haploid number e.g. 4n=92
16
Q
What is meant by the term “Aneuploid”?
A
- Any chromosome number which is not an exact multiple of haploid number - due to extra or missing chromosome(s) e.g. 2n+1=47
17
Q
What are the different types of numerical chromosomal abnormalities?
A
- Trisomy - Type of aneuploidy in which there are three instances of a particular chromosome, instead of the normal two
- Monosomy - Type of aneploidy in which there is only one instance of a particular chromosome
-
Mosaicism - When a person has 2 or more populations of cells with a different number of chromosomes
- E.g. person may have a population of haploid cells (46 chromosomes) and another population of cells with aneuploidy
18
Q
Give a brief overview of Meiosis
A
- DNA is replicated so each chromosome has 2 sister chromatids
- Recombination occurs between homologous chromosomes
- Meiosis I: homologous chromosomes line up next to each other at the equator of the cell; get attached to the mitotic spindle and then get separated to opposite spindle poles
- Now each of the 2 cells has 23 pairs of chromosomes (diploid)
- Meiosis II: sister chromatids line up at the equator of the cell and get attached to the mitotic spindle and get separated to opposite spindle poles
- Now each of the 4 daughter cells only has 23 chromosomes (haploid)
19
Q
Give a brief overview of Mitosis
A
- Prophase: Chromosomes condense, centrosomes move to opposite poles, mitotic spindle forms
- Prometaphase: Breakdown of nuclear envelope, chromosomes attach to mitotic spindle
- Metaphase: Centrosomes are at opposite poles, Homologous chromosomes line up one behind the other at the equator of the mitotic spindle and get attached to mitotic spindle
- Anaphase: Sister chromatids separated to opposite spindle poles
-
Telophase: chromosomes decondense, nuclear envelope reforms
- Now each of the 2 cells has 23 pairs of chromosomes (diploid)
20
Q
What is it called when chromosomes/chromatids are pulled to opposite ends of the cell during anaphase?
A
- Disjunction
21
Q
How does aneuploidy arise?
A
- Primary mechanism by which aneuploidy arises is non-disjunction - when homologous chromosomes DON’T separate from one another
22
Q
What happens if non-disjunction occurs during meiosis I?
A
- If non-disjunction occur during meiosis I both copies of a pair of homologous chromosomes will end up in one daughter cell while the other daughter cell doesn’t get any
- Meiosis II will occur as normal so daughter gametes formed from cell that got both pairs of homologous chromosomes will end up with 2 copies of that particular chromosome (disomic)
- Daughter gametes formed from cell that didn’t get either pair of the homologous chromosomes don’t have any copies of that particular chromosomes (nullisomic)
23
Q
What happens if non-disjunction occurs during meiosis II?
A
- If non-disjunction occurs during meiosis II then both sister chromatids of a particular chromosome end up in one daugther gamete while the other daughter gamete doesn’t get any
- This means one daughter gamete is disomic with respect to that chromosome while the other is nullisomic
24
Q
Give some examples of the most common autosomal trisomies
A
- Trisomy 21
- Trisomy 18 (Edward’s syndrome)
- Trisomy 13 (Patau Syndrome)
25
What happens if non-disjunction occurs during mitosis (Mitotic non-disjunction)
* Initially results in one cell have 3 copies of a particular chromosome (trisomic) and one cell only having one copy of that same chromosome (monosomic)
* All monosomic cells are broken down
* Trisomic cells are maintained
* So end result is that you have some trisomic cells, some disomic cells as mitotic disjunction doesn't occur in every cell and monosomic cells
* This is called mosaicism
26
What are the 2 mechanisms that result in mosaicism?
* **Post-zygotic nondisjunction or mitotic non-disjunction:** All diploid cells (2n) become a mixture of diploid (2n) and trisomic (2n+1) cells
* **Anaphase lag (trisomic rescue):** Meiotic non-disfunction results in all cells being trisomic for a particular chromosome
* Every one of those trisomic cells goes through mitosis but during anaphase one of the 3 copies of that particular chromosome doesn't connect to the mitotic spindle
* This results in a micronucleus forming around that chromosome which means it doesn't end up in any of the daughter cells
* All daugther cells where anaphase lag occured are disomic (2n) and daughter cells where anaphase lag hasn't lagged are still trisomic (2n+1)
27
What is the clinical relevance of mosaicism?
* Mosaic phenotype is thought to be less severe
* However, it's difficult to assess:
* What proportion of cells will be trisomic/disomic
* What tissues/organs will be affected as a result of mosaicism
28
What are some examples of conditions caused by mosiacism in gametes?
* Turner's syndrome - (46, XX / 45, XO)
* Klinefelter's syndrome - (46, XY / 47, XXY)
29
How common are autosomal and gamete monosomies?
* Autosomal monosomies are very very rare
* Monosomies in gametes more common. e.g. Turner's syndrome
30
What is the difference between full monosomy and partial monosomy?
* **Full monosomy** - Arises via meiotic non-disjunction
* **Partial monosomy (microdeletion syndromes) **- Far more common and mechanism different
31
What are all the different combinations of gametes that result in trisomic/monsomic zygote conditions
* **Nullisomic gametes**
* O + X chr = XO = Turner’s (physically female)
* O + Y chr = lethal
* **Disomic gametes (XX)**
* XX + X chr = XXX = Triple X syndrome
* XX + Y chr = XXY = Klinefelter’s (physically male)
* **Disomic gametes (XY)**
* XY + X chr = XXY = Klinefelter’s
* XY + Y chr = XYY = XYY syndrome
32
How do you generate a karyotype from a patient to determine if there are any abnormalities?
* Entire process takes about 3 days
33
How can you analyse a developing foetus to see if it has any chromosomal abnormalities (prenatal diagnosis)?
* Chorionic Villus Sampling
* Amniocentesis
34
Give an overview of Chorionic Villus Sampling
* Transvaginal or transabdominal injection used to take sample of chorionic villus which is then analysed using karyotyping/FISH/QF-PCR
* Carried out at 11-14 weeks
* Miscarriage rate of 0.5% to 1%
* May also cause:
* Maternal contamination (getting maternal cells in sample)
* Transverse limb defects
35
Give an overview of Amniocentesis
* Injection used to take sample of amniotic fluid which is then analysed karyotyping/FISH/QF-PCR
* Carried out anytime after 16 weeks
* Miscarriage rate of 0.5-1%
36
What is G-banding?
* G-banding (Giemsa banding) involves staining chromosomes with giemsa stain
* Allows chromosomes to be viewed during metaphase
* Chromosomes are Lined-up based on:
* **Size**
* **Banding**
* **Centromere position**
* Dark bands = Heterochromatin
* Light bands = Euchromatin
37
Why do chromosomes viewed using G-banding have a badning pattern?
* Because chromosomes contain both euchromatin and heterochromatin and both stain differently when using the Giemsa stain
* **Euchromatin** = GC-rich; loosely packed; genes active
* **Heterochromatin** = AT-rich; tightly packed; genes inactive
38
Give a brief overview of FISH and explain how it works
* FISH = Fluorescent in situ hybridisation
* Uses cultured cells
* Looks at metaphase spread of chromosomes (same as karyotyping)
* Can detect abnormalities of up to 5-10Mb
1. Design Fluorescent probe to a chromosomal region of interest
2. Denature probe and target DNA
3. Mix probe and target DNA together (hybridisation)
4. Probe binds to the target DNA on the chromosome of interest
5. Target fluoresces or lights up
39
What is QF-PCR and why is quicker to perform than FISH or karyotyping?
* QF-PCR (Quantitative fluorescence PCR) is a technique that uses fluorescent probes to amplify and then bind to specific microsatellites that are known to be on a specific chromosome
* QF-PCR is used to detect chromosomal abnormalities (normally trisomies) by quantifying the no. of a specific microsatellite
* E.G. If probes detect microsatellite X, known to be on chromosome 21, 3 times then you know it's trisomy
* QF-PCR is quicker than FISH/Karyotyping because you don't need to use cultured cells
40
How can you analyse a developing foetus to see if it has any chromosomal abnormalities, non-ivasively (Non-invasive diagnosis)?
* **Cell-free foetal DNA (cffDNA)** – DNA fragments in maternal plasma from 10 wks onwards
* They can be extracted and isolated from maternal blood and be tested to see if the foetus has any abnormalities
* For monogenic disorders you use PCR to amplify particular gene in foetal DNA which may cause particular disease before testing that gene
* For aneuploidies you use NGS to test foetal DNA
* **SAFE TEST** - Uses these principals to test for Trisomies 13, 18, 21 - these still do need to be confirmed using amniocentisis/CVS
