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Flashcards in Test 2 Deck (80)
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1
Q

Monogenic definition

A

strong genetic influence by 1

gene

2
Q

Recessive definition

A

Need two defective copies to be

affected

3
Q

Cystic fibrosis is due to:

A

mutations in the gene for the CF transmembrane conductance regulator
(CFTR) gene, Cl- channel

4
Q

Characteristics of cystic fibrosis:

A
  • fatal monogenic recessive disease
  • congenital (present at birth)
  • highest frequency in Northen Europe
  • Heterozygote/carrier frequency of 1/20
  • General frequency in Caucasian populations: 1/1600-3000
  • less common in African, Arab or Asian populations
5
Q

Inheritance pattern of CF with two carriers as parents:

A
  • 25% offspring unaffected, 50% carriers (unaffected), 25% have the disease
6
Q

Inheritance pattern of CF with one normal parent and one parent as a carrier

A
  • 50% of offspring are carriers (unaffected), 50% of offspring do not carry the mutation at all (unaffected)
7
Q

Phenotype/clinical presentation of CF:

A

– increased sweat Na and Cl
– pancreatic insufficiency
– pulmonary infections

  • thick mucous production
  • 90% of males are infetile - congenital bilateral absence of the vas deferens
  • > 90% of deaths due to recurrent infection, most commonly with Pseudomonas aeruginosa
8
Q

Root of pathophisology of CF

A

Stems from a defect in CFTR, a
chloride transporter on cell
membrane

9
Q

Pathophysiology of CF

A
  • less chloride channel activity
  • change in regulation of epithelial sodium channel
  • increased sodium, increased water absorption, decreased water content of secretions (also increased sodium in sweat)
  • mucous becomes thicker -> impaired flow, plugs and obstructions
10
Q

Effect of pathophysiology of CF on GIT

A
  • Blockage of pancreatic ducts.
  • Loss of water from GIT
  • Meconium ileus
  • Constipation
  • Rectal prolapse
11
Q

Effect of pathophysiology of CF on lungs

A

Decreased periciliary volume + thick

mucus leads to poor ciliary clearance of pathogens and recurrent infections

12
Q

CFTR gene

A
  • Chromosome 7q31-32 - 250Kb region
  • 27 exons, 6.5Kb transcript
  • Regulated by cAMP sensitive protein kinase
13
Q

CFTR protein

A
  • A 1480 aa membrane protein.
  • A Cl- ion channel expressed in the apical membrane of exocrine
    epithelial cells.
  • Member of the ATP binding cassette family of transporters
14
Q

Mutations in CTFR

A
  • > 1,000 mutations described.
  • One major mutation in Caucasians, ΔF508, accounts for ~ 70%
  • A few at frequency of 1-3%; the rest at very low frequency.
15
Q

Classic CF clinical phenotype is associated with:

A
  • Pancreatic exocrine insufficiency (highest correlation between genotype and phenotype)
  • COPD (lowest correlation between genotype and phenotype for lung manifestations)
  • Abnormal concentrations of sweat electrolytes
  • No vas deferens in males
  • Bowel obstruction
16
Q

Pulmonary manifestations in genotype/phenotype correlations (CF)

A
  • Most common cause of morbidity and mortality.
  • But there is a poor correlation with severity of genotype and the age on
    onset, severity and progression.
  • For example, even among homozygotes for the most common mutation
    (DF508), lung function may vary from normal to severe dysfunction
17
Q

How heterozygous CF patients have an advantage against diarrhoea

A
  • in normal patients: toxins released by cholera and E coli act on CFTR, cause increased fluid flow in intestine –> diarrhoea
  • CF homozygotes don’t secrete chloride ions in response to bacteria
  • Mutations can protect against diarrhoea
18
Q

How heterozygous CF patients have an advantage against typhoid fever

A
  • Salmonella typhi enter epithelial cells via interactions with CFTR.
  • One study, showed an 86% reduction in internalization of S. typhi into the gastrointestinal tract, of mice
    heterozygous for Cftr-ΔF508, relative to that in wild-type mice
  • Thus, selection for typhoid fever resistance provides another possible
    explanation for the high frequency of CFTR mutations
19
Q

Newborn screening for CF

A
  • Measure immunoreactive trypsinogen (IRT), 48-72h after birth (these levels are raised in CF), blood is from heel prick
  • Genotyping: two mutations = CF, no mutations = unlikely to have CF, one mutation = carry out sweat test to determine electrolytes
  • CF patients cannot absorb NaCl from sweat
20
Q

Sweat Test Interpretation for CF

A
  • Australian recommended values for sweat chloride in infants in newborn screening (should be done after 1st postnatal week):
  • Cl 60 mmol/l, cystic fibrosis;
  • Cl 30–59 mmol/l, borderline;
  • Cl 29 mmol/l, normal.
  • Up to 25% of infants with MI do not have an IRT value above the cutoff level. Any neonate with a family history of CF or Meconium Ileus SHOULD be followed up
21
Q

Fragile X syndrome:

A
  • most common inherited intellectual disability
  • can range from learning disabilities to severe cognitive disabilities to intellectual disabilities
  • 30% of individuals with Fragile X have autism
  • 2-6% of individuals with autism have Fragile X
  • diagnosed with bloods or DNA testing
22
Q

Prevalence of Fragile X

A
  • Affects 1/4000 males and 1/6000-8000 females
  • Can appear in all socio-economic backgrounds
  • 50% of females with full mutations have some form of ID
23
Q

Clinical features of Fragile X

A
  • Large ears, long narrow face w/ prominent forehead, mitral valve prolapse, seizures, eye problems
  • FGX mutations affecting brain development - mental impairment, developmental delays, learning disabilities
24
Q

Cause of Fragile X disorder

A
  • more than 200 repeats in CGG expansion
  • this leads to hypermethylation of cytosine residues
  • causes deactivation of FMR1 gene (no FMRP produced in Fragile X)
25
Q

Analysis of mutations of Fragile X

A
  • Stable: <45 repeats, unmethylated, individuals not affected
  • Grey zone: 45-54 repeats, unmethylated, individuals not affected
  • Pre-mutation: 55-200 repeats, unmethylated, individuals usually not affected
  • Full mutation: >200 repeats, completely methylated, 50% of women and 100% of men affected
26
Q

Fragile X associated primary ovarian insufficiency

A
  • carriers of FX pre-mutation (55-200 CGG repeats)
  • produce more mRNA but less FMRP protein
  • in females, menopause is 5 years earlier and 23% of female individuals have ovarian insufficiency
  • toxic RNA gain is said to be responsible
27
Q

Fragile X associated tremor ataxia syndrome

A
  • affects older adults that carry pre-mutation (increased rate with age, increased rate of hypertension)
  • intention tremor and cerebellar gait ataxia are main clinical features
  • affects 40% of male carriers and 8-16% female carriers
28
Q

FMR1 gene

A
  • chromosome Xq27
  • CGG repeats happen in 5’-UTR (healthy individuals have 6-55 copies)
  • methylation of CpG site -> silencing gene -> deficiency in FMRP protein and intellectual disability
29
Q

Lab testing for Fragile X

A
  • PCR, Southern Blot -> no. of CGG expansions
  • Microarray and MLPA to detect deletions
  • Next Gen sequencing
30
Q

Fragile X treatment

A
  • Glutamate antagonists - reduce activity of metabotropic receptor
  • Lithium - improved behaviour but not cognition
  • MMP9 - reversal of structural damage in neurons
  • GABA agonists (arbaclofen) - alleviate anxiety and learning disabilities
31
Q

Clinical features of Huntington’s disease

A
  • Progressive neurodegenerative disease leading to dementia
  • Symptoms include mood and character changes, defects in memory and attention, progressing to movement disorders
  • Onset is usually at 35-55 years and symptoms evolve over 12-15 years
32
Q

Genotype/phenotype of Huntington’s

A
  • Autosomal dominant
  • Instability of CAG repeat length in sperm and oocyte DNA
  • A parent that has Huntington’s (heterozygous) has a 50% chance of passing it to their children if they are with an unaffected partner
33
Q

Genomic information HD gene (Huntington’s)

A
  • On chromosome 4p16
  • has 67 exons
  • protein product ins huntingtin
  • mRNA codes for 3144 amino acids
34
Q

Errors in HD gene (Huntington’s)

A
  • repeat region of CAG (197-265bp)
  • codes for glutamine
  • 23 glutamines in repeating sequence of the normal protein
35
Q

HD assay (Huntington’s)

A
  • Determine CAG repeat sequence
  • Use sizing standards between 27-40 repeats
  • Participate in proficiency testing
  • Resolve homozygous alleles
36
Q

Huntington’s genome guidelines:

A
  • Normal allele: <26 CAG repeats, normal phenotype
  • Mutable normal allele: 27-35 CAG repeats, normal phenotype (risk that children will develop it)
  • HD allele (1): 36-39 CAG repeats, HD phenotype but may not develop it
  • HD allele (2): >40 CAG repeats, HD phenotype
37
Q

Genetic testing for huntingtons

A
  • predicitive testing - future onset
  • diagnostic testing - those who develop symptoms
  • Prenatal: CVS-DNA, direct gene analysis, exclusion testing
38
Q

Pathology (Huntington’s)

A
  • exact function of Huntingtin protein is unknown, but accumulation of abnormal protein can cause neurological changes and interferes with neurotransmitters
39
Q

Possible mechanism of action (Huntington’s)

A
  • Huntintin widely expressed in cytoplasm
  • amino-terminal fragments of mutant huntingtin aggregate in nuclear and cytoplasmic inclusions and destruct neurons
  • primary cause: soluble or aggregated form of mutant Huntingtin
40
Q

Huntington’s disease can also be involved with:

A
  • breast cancer
  • hereditary haemachromatosis
  • familial hypercholesterolaemia
41
Q

Principles of predicitive testing (Huntington’s):

A
  • information collected, at risk individual is identified
  • accurate risk information is given
  • implications are explored and patient is provided with a time out to consider
  • further counselling and details of results session
  • informed consent to collect samples
  • results and follow up
42
Q

Five domains to consider when diagnosing Huntington’s

A
  • Occupational
  • Fiscal
  • Activities of daily living
  • Household chores
  • Residence
43
Q

Forensics in the past

A
  • ABO blood groups to identify people
  • Not sensitive, lots of biomaterial
  • Low discrimination, exclusionary, supporting evidence
  • Cannot ID unknown suspects, no databasing
44
Q

Forensics now

A
  • DNA to identify people
  • Very sensitive, invisible traces, many crimes/cold case
  • High discrimination, evidentiary, primary evidence
  • Can ID unknown suspects, amenable to databasing
45
Q

Services that the forensics lab provide:

A
  • DNA analysis (criminal, DVI, parentage, missing persons, databases)
  • Hair analysis, animal species identification, stain identification
  • Fabric damage assessment, blood stain pattern analysis
46
Q

Types of offences - serious crime

A
  • homicides
  • sexual assaults
  • armed robberies
  • assault/wounding
  • drug possession or manufacture
  • criminal parentage
47
Q

TYpes of offences - volume crime

A
  • burglary
  • break and enter
  • motor vehicle theft
48
Q

Types of offences - other

A
  • ID of unknown deceased
  • missing persons
  • disaster victim identification (natural disaster, accident, criminal action)
49
Q

Stages of DNA profiling:

A
  • Extraction
  • Quantitation
  • Amplification
  • Analysis
50
Q

DNA Extraction (stages of DNA profiling)

A
  • various methods: manual and automated
  • inhibitory substances common
  • complex substrates
  • liquid handling robotics
51
Q

DNA Quantitation (stages of DNA profiling)

A
  • Real-time PCR

- Human-specific

52
Q

Amplification (stages of DNA profiling)

A
  • Multiplex PCR

- thermal cyclers

53
Q

Analysis (stages of DNA profiling)

A
  • Capillary Electrophoresis (rapid, accurate, sensitive)
54
Q

Short tandem repeats

A
  • high heterozygosity (differentiate people)
  • regular repeat unit (predictable alleles)
  • distinguishable alleles
  • robust amplification
  • can be multiplexed (less DNA required, faster)
  • small product sizes (<500 bp range) are better for degraded DNA
55
Q

ABO blood typing vs Forensic DNA profiling

A
  • A blood group can be present in 47% of the population, and is better at excluding people rather than including people
  • The chance of two unrelated people have the same DNA profile is less than 1 in 100 billion, and the world population is around 7.7 billion –> good at individualising people
56
Q

When DNA profile statistics don’t matter

A
  • At trial, it usually doesn’t matter whose DNA it is, it matters how it got there
57
Q

What you can and can’t determine from a forensic DNA profile

A
  • you can determine the gender of the individual

- can’t determine ethnicity, racial background, age, physical appearance or illnesses

58
Q

Negative results in a forensics laboratory

A
  • you can only comment on the findings in the laboratory, not if an event had occured or not
  • sperm is never detected in 50% of sexual assault cases
59
Q

DNA Databases state and national

A

WA: Criminal Investigation (made July 2002)
National: NCIDD (National Criminal Investigation DNA Database), ACIC (Australian Criminal Intelligence Commission)

60
Q

Markers used in fornesic biology

A
  • ABO blood groups: low discrimination, fast analysis
  • RFLP multi locus probes: high discrimination, slow analysis
  • Multiplex STRs: high discrimination and fast analysis
61
Q

What STRs and SNPs detect in Next Gen and Massively Parallel

A

The identity of the individual:
- Global autosomal STRs, Y-STRs, X-STRs, Identity SNPs

What they look like:
- Phenotypic SNPs

Where they are from:
- Biogeographical Ancestry SNPs

62
Q

Examples of some genealogy sites

A
  • LivingDNA
  • MyHeritage
  • FamilySearch
  • Ancestry
  • Family Tree DNA
  • 23 and Me
63
Q

Myths - the CSI effect

A
  • There is always forensic evidence
  • Evidence is easy to locate
  • Evidence is easy and quick to test
  • Evidence always provides simple, clear-cut answers
  • Cases are always solved quickly
64
Q

Intrinsic pathway to common pathway of coagulation

A
  • XII converts to XIIa
  • XIIa converts XI to XIa
  • XIa converts IX to IXa, which binds to VII and protein C
  • Compound allows conversion of X to Xa, which binds to Va
  • This compound converts prothrombin (II) to thrombin
  • thrombin converts fibrinogen (I) to fibrin
65
Q

Extrinsic pathway to common pathway of coagulation

A
  • tissue factor (III) binds to VIIa
  • compund activates conversion of X to Xa, which binds to Va
  • This compound converts prothrombin (II) to thrombin
  • thrombin converts fibrinogen (I) to fibrin
66
Q

Factor V East Texas severity:

A
  • a moderately severe

bleeding disorder

67
Q

Factor V East Texas symptoms:

A
  • Easy bruising
  • Epistaxis
  • Bleeding after trauma or surgery
  • Menorrhagia
68
Q

Lab characteristics Factor V East Texas

A
  • prolonged PT and aPTT
  • Normal factor levels
  • Mixing studies do not suggest presence of an inhibitor
  • Patient is not on a pharmacological inhibitor
  • [Platelet] = Normal
69
Q

Hypothesised principle of FV East Texas

A
  • novel mutation of F5
  • A→G in Exon 13 F5
  • Ser→Gly at a.a. 756
  • [FV] is normal in affected individuals
70
Q

Location of F5 mutation

A
  • AA 756 is in the B domain
  • Only the Heavy and Light Chains are required for factor V procoagulant activity
  • The B-domain is not required for factor V procoagulant activity
71
Q

Cleavage of Factor V

A
  • Factor V is activated by thrombin or factor Xa. Thrombin cleaves the B-domain at aa’s 709, 1018, and 1545.
  • Factor Xa cleaves the B-domain at aa’s 709 and 1018, but is very inefficient at
    cleaving at aa 1545
  • Both proteins separate the basic region from the acidic region, producing an active form of factor V
  • Factor Xa is in the B domain near the acidic region, thrombin is in the heavy chain
72
Q

Function of TFPI

A
  • TFPI inhibits fXa and TF:fVIIa
  • At normal physiological concentrations, TFPI must bind to fXa before it can inhibit TF:fVIIa
  • At elevated concentrations, TFPI can inhibit TF:fVIIa
    independently of fXa
  • TFPI binds to forms of fVa that retain the acidic region
73
Q

How TFPIa binds FV

A
  • Basic region of TFPIα binds to acidic region of fV - the basic regions of each factor are homologous
74
Q

Exon 13 FV East Texas

A
  • Exon 13 is shortened
  • PCR performed using F primer in exon 12 and R primer in exon 14
  • Unaffected - complete exon 13 transcript (top) is bright
  • Affected: novel exon 13 transcript (bottom) is bright
75
Q

How exon 13 is shrotedin FVET

A
  • The A→G mutation produces a novel splicing donor site that causes an in-frame deletion of 702 aa
76
Q

Xa- activated Factor Va vs fV-Short in FVET

A
  • Xa- activated Factor V - B domain is from aa 1019 to 1545

- fV-Short - B domain is from aa 1458 to 1545

77
Q

Calibrated automated thrombography FV East Texas

A
  • measures thrombin generation in plasma
  • plasma incubated with low TF concentration
  • Coagulation is initiated with the addition of Ca2+
  • Thrombin generation is lower in FV East Texas patients
78
Q

Relationship between TFPIa and TG

A

↑ Plasma TFPIα is the cause of ↓ TG

79
Q

What type of mutation is FVET

A

indirect, gain of function

mutation

80
Q

Individuals with Fv Amsterdam:

A
  • have ↓ TG that is corrected by anti-TFPI antibody
  • have a shortened fV protein
  • due to a novel mutation in exon 13 of FV with deletion of aa 623 in the B domain (basic region removed)