MCBHD Flashcards
What is meant by autosomal dominant?
- manifests in HETEROZYGOUS form
- multiple generations affected
- male to male transmission
- 50% risk to offspring
- only need a mutation in one copy of the gene to manifest symptoms
What are characteristics of autosomal inheritance patterns?
- most individuals have an affected parent (not everyone because of cases of new mutations or incomplete penetrance)
- males and females are equally likely to inherit the allele and be affected
- risk for each child of an affected parent is 0.5
- if an affected individual’s siblings/children are not affected, and they do not carry the mutation they cannot pass it on to their own offspring
What is the definition of PENETRANCE?
The percentage of individuals expressing the disorder to any degree (sever or mild) - many dominant disorders show age dependent penetrance
What is EXPRESSIVITY?
Variation in the severity if a disorder tween individuals with the same mutation
What is the new mutation rate?
‘De novo’ mutation rate varies considerably between different AD conditions
What is reproductive fitness?
In some AD disorders mutations carriers do not reproduce - the disorder is maintained in the population by new mutations
What is somatic mosaicism?
A new mutation arising at an early state in embryogenesis - present only in some tissues/cells
What is germ-line mosaicism?
(Gonadal mosaicism) a new mutation arises during oogenesis or spermatogenesis - the mutation is present in a variable proportion of the gametes and can be transmitted to the offspring
What is paternal age effect?
The chance of a new mutation increases with advancing paternal age
What is ANTICIPATION?
Worsening of disease severity in successive generations - characteristically occurs in triplet repeat disorders
What is AUTOSOMAL RECESSIVE inheritance?
- manifests in homozygous/compound heterozygous form
- carriers are not affected
- usually one generation affected
- May be consanguinity - eg cousin marriages
- need to have mutations in both copies to be affected
- carriers have a normal copy which is sufficient to prevent symptoms
- unless it is a common disorder, or there is consanguinity, normally only a single generation is affected
What are characteristic patterns of autosomal excessive inheritance?
- males and females are equally likely to be affected
- the trait is often found in clusters of siblings but not in their parents and offspring
- the more rare a trait in the general population, the greater ten chance it was a consanguineous mating
- recurrence risk is 1/4 for each sibling of an affected person
- carrier probability of 2/3 of normal siblings of an affected person
- all offspring of an affected person are OBLIGATE CARRIERS
About x-linked inheritance…
Women have 2 X chromosomes –> two copies of x-linked genes
Men have 1 X and 1 Y –> only a single copy of x-linked genes
Can be…
RECESSIVE: women are carriers, no make to make transmission
DOMINANT: women are affected, males more severely affected/lethal
What are patterns of x-linked inheritance?
- x-linked genes are never passed from father to son
- males are never carriers
- makes are more likely to be affected because they only have one copy to the gene
- affected males get the disease from their mothers (or are new mutations)
- all of the daughters of affected males are obligate carriers
- children of carrier females have a 50% chance of receiving the mutant allele
What is skewed x-inactivation?
Normally the majority of genes on one of a woman’s x-chromosomes are inactivated. This is generally random but ~10% of women have uneven or skewed x-inactivation >80:20%
What are manifestation carriers?
Some women do have symptoms in x-linked recessive conditions eg cardiomyopathy in DMD - unfavourable skewing of x-inactivation may help to explain this
About mitochondrial inheritance…
- rare
- small circular molecule
- exclusively maternally inherited/transmitted
- there may be:
Homoplasmy: only one type of mtDNA
Heteroplasmy: more than one type of mtDNA
Threshold effect: normal mitochondrial function below a proportion of abnormal mtDNA but abnormal above it
What is heteropasmy?
The presence of more than one type of organelles genome (mtDNA) within a cell of individual. It is an important factor in considering the severity of mitochondrial diseases.
What is the basics if mitosis and meiosis?
Daughter cells created after mitosis are generally identical to parent cells.
Meiosis is the type of cell division by which eggs and sperm are produced. Meiosis involves a reduction in the amount of genetic material
- comprises two successive nuclear divisions with only one round of DNA replication
- one parent cell produces four daughter cells
- daughter cells have half the number of chromosomes found in the original parent cell and with crossing over, are genetically different.
How do we look at chromosomes?
- blood same or CVS/amnio
- culture cells (lymphocytes)
- arrest cells in metaphase
- burst cells to release chromosomes
- mount on a slide and stain
What are methods of chromosome identification?
- G-banded ideogram: giemsa stain, most common
- FISH
- array - comparative genome hybridisation (Array-CGH)
What is FISH?
Fluorescence in situ hybridisation
Specific DNA probe used to identify a region if interest (gene, locus, centromere, etc)
cultured cells, metaphase spread
microscopic (5-10 Mb)
What is aneuploidy?
HAPLOID: one set of chromosomes (n=23) as in a normal gene
DIPLOID: cell contains two sets of chromosomes (normal in human)
POLYPLOID: multiple of the haploid number
ANEUPLOID: chromosome number which is not an exact multiple if the haploid number - due to extra or missing chromosome(s)
Changes can be in the germline or be as a result of mosaicism
What is mosaicism?
The presence of two or more genetically different cell lines derived for a single zygote
About Down’s syndrome
Trisomy 21 95% patients NDJ (usually maternal meiosis I) 5% robertsonian chromosome ~2% mosaic 'Order egg model' - maternal age effect
What are the clinical features of Down’s syndrome?
1 in 650-1000 live births
Most common cause of mental retardation
Hypotonia, particularly in newborn period
Developmental delay
Cardiac abnormalities; AV canal defects most common
GI abnormalities; duodenal stenosis/atresia, imperforate anus, Hischsprung disease
ALL/AML - 10-20 x relative risk
Conductive hearing loss
Features of Alzheimer’s >40 years
About Edwards syndrome…
Trisomy 18 Incidence ~1 in 6000 live births Intrauterine growth retardation Micrognathia Cleft lip +\- palate Short palpebral fissures Fixed flex ion deformities of fingers Heart defect >95% - VSD/ASD/PDA Inguinal/diaphragmatic hernias Renal malformations
About Patau syndrome…
Trisomy 13 - primary or secondary robertsonian translocation
~1 in 10,000 live births
Midline defects: hyotelorism, holoprosencephaly,midline cleft lip/palate, scalp defect
Post axial polydactyly
Heart defects/renal abnormalities
Survival - most die by 1 month
What are the basic rules of pedigree drawing?
- males are squares
- females are circles
- partners have a line between them
- siblings have a line above them
- there is a line down for children
- affected people are shaded in
- carriers have dots in
About turner syndrome…
- incidence 1/4000 female births
- 45X0 or mosaic (45X0/46XX, 45 XO/47XXX, 45X0/46XY)
- > 60% miscarry
- high incidence in spontaneous abortions (9-10%)
- raised at nuchal translucency/cystic hygroma
- at birth: oedema of hands and feet, neck webbing, coarctation of aorta, renal malformation
- short stature
- infertility secondary to gonadal dysgenesis
- intellectually normal
What is turner mosaicism?
- 45X/46XY mosaicism - usually normal male (90%), but with potential for gonadoblastoma
- mosaicism with ring or isochromosome Xq results in variant Turner syndrome
What is Klinefelter syndrome?
- 47XXY
- incidence 1/1000 male live births
- phenotype mild and variable - some cases undetected
- Barr body present
- NDJ paternal meiosis I (50%), others NDJ maternal meiosis or zygotic mitotic error (mosaic)
- variants 48,XXYY, 48,XXXY etc
What are clinical symptoms of Klinefelter syndrome?
- may present prenatally, during childhood with behavioural problems, or adulthood with infertility
- tall stature
- eunuchoid body habitus
- some behavioural and minor learning difficulties
- lack of secondary sexual characteristics - treat with testosterone
- infertility
About structural rearrangements…
Approximately 1/400 newborns have a structural rearrangement
BALANCED: no net gain or loss of genetic material
UNBALANCED: net gain or loss of genetic material
RECIPROCAL or ROBERTSONIAN
usually no deleterious phenotype unless breakpoint affects regulation of a gene
What is reciprocal translocation?
- two afrocentric chromosomes join near centromere with the loss of p arms
- balanced carrier has 45 chromosomes
- if 46 chromosomes present including Robertsonian then must be unbalanced
- P arms encode rRNA (multiple copies so not deleterious to lose some)
- 13q14q and 14q21q relatively common
About deletions…
- breakage and loss of eccentric fragment
- terminal or interstitial
- monosomic region - haploinsufficiency of some genes
- phenotype usually abnormal and specific for size and place of deletion
- 1/7000 live births
About microdeletions/duplications…
- many patients has no abnormality visible on metaphase spread
- high resolution banding, FISH and now CGH showed ‘micro’ deletions
- only a few genes may be lost or gained - contiguous gene syndrome
- areas of low copy number repeat sequences - recombination error?
What happens in a cytogenetics laboratory?
chromosome analysis
pre and post-natal testing, adult testing: foetal blood, amniotic fluid, CVS biopsy, solid tissue; venous blood, tumour tissue
WHY?: ambiguous genetalia/indeterminate gender, known familial chromosome rearrangement, multiple miscarriages
HOW?: G-banding, FISH, QF-PCR, array-CGH
About G-banding…
- giemsa stain
- metaphase
- line up based on size, banding and centromere position
uses a chemical stain, uses metaphase chromosomes, takes several days at least, looks for aneuploidies, translocations and very large deletions
Why do you get chromosome banding?
Chromatin: 2 different sorts: euchromatin and heterochromatin, euchromatin = GC-rich; loosely packed; genes active, heterochromatin = AT-rich; tightly packed; genes inactive, stain differently
what are the steps of FISH testing?
- fluorescent probe
- denature probe and target DNA
- mix probe and target DNA
- probe binds to target
What are examples of FISH tests?
- 22q deletion syndrome
- prader-willi syndrome (chromosome 15)
- cri-du-chat (chromosome 5)
You have to know what causes a disease in order to be able to test for it by standard FISH, because you have to be able to design a specific probe.
What is QF-PCR?
- quantitative fluorescence PCR
- triremes 13, 18 and 21
- uses microsatellites
- looks at how many copies of a chromosome the patient has
- uses fluorescent probes for specific micro satellite markers on specific chromosomes
- uses extracted DNA
- quick (~48 hours)
- looks for aneuploidies
- need to know what you’re looking for
What are micro satellites?
di, tri, tetra, penta, hexa nucleotide sequence with a variable number of repeats
- number of repeats is constant within an individual
- number of repeats varies between individuals
What is array-CGH?
- comparative genomic hybridisation
- sub-microscopic chromosome abnormalities (~60kb)
- micro deletions, micro duplications, CNVs
- main indication = developmental problems, dysmorphia
- uses fluorescent probes to differentiate between patient and control
- uses extracted DNA
- looks for deletions and duplications
What are Copy Number Variations (CNVs)?
- deletions/duplications
- 10kb-5000kb
- can protect e.g. from HIV
- can be detrimental e.g. autism and schizophrenia
- many do nothing
- ~12% of the genome = CNV
What is the process of array-CGH?
- extract DNA
- test DNA labelled with fluorescent dye
- control DNA labelled with different fluorescent die
- mix DNA tighter
- apply denatured DNA to array –> hybridise
- measure relative fluorescence
What is the sequence of sanger sequencing?
- DNA isolation and amplification (generally each exon separately)
- denature
- anneal primer
- extend
- terminate
- denature
- read; sort fragments and determine sequence
DNA fragments are drawn through the capillary; the smaller fragments pass through more quickly than larger fragments
What is NextGen sequencing?
- targeted
- whole exome
- whole genome
Looks at each base and identifies small substitutions and small indels. It can look at ALL the genes at once and uses different chemistry from Sanger sequencing.
