mutations and genetic analysis Flashcards
defne trisomy
3rd copy on one of the chromosomes
define monosomy X
only one X chromosome
define tetraploidy
whole extra set of chromosomes
numerical chromosome abnormalities
change in the number of chromosomes
structural chromosome abnormalities
large change in the structure of a chromosome
mutational chromosome abnormalities
changes to the DNA sequence
origins of chromosome abnormalities - non-disjunction
the failure of one or more pairs of homologous chromosomes or sister chromatids to separate normally during nuclear division, usually resulting in an abnormal distribution of chromosomes in the daughter nuclei.
where do non-disjunction abnormalities tend to occur
maternal meiosis
origin of non-disjunction
In the female meiosis, the cells are in meiosis I for a very long time before they separate and mature into eggs - this can explain why non-disjunction problems are more common in maternal meiosis
define aneuploidy
the condition of having an abnormal number of chromosomes in a haploid set.
autosomal aneuploidy syndromes
trisomy 21 (down's syndrome) trisomy 13 (Patau syndrome) trisomy 18 (Edward's syndrome)
trisomy 21
Down's syndrome Incidence increases with advancing maternal age • Characteristic facial dysmorphologies • IQ less than 50 • Average life expectancy (50-60 years) • Alzheimer’s disease in later life
chromosomal findings in Down’s syndrome
○ Trisomy 21: non-disjunction (95%), usually maternal origin
○ Unbalanced Robertsonian translocation (4%)
○ Mosaicism (1%) - some of the cells in the body have trisomy 21 but others are normal, thought to arise due to non-disjunction error during early cell division
Trisomy 13
Patau syndrome
• Multiple dysmorphic features and mental retardation
• About 5% die within first month, very few survive beyond first year
• Non-disjunction (90%), maternal origin
• Unbalanced Robertsonian translocation (10%)
trisomy 18
Edwards syndrome
• Severe developmental problems (but is more variable); most patients die within first year, many within first month
Non-disjunction (90%), maternal origin
sex chromosomes aneuploidy syndromes
45, X - turner syndrome
47, XXY- Klinefelter syndrome
45, X
Turner syndrome
• Incidence at conception much greater, about 97% result in spontaneous loss
• Females of short stature and infertile
○ No Y chromosome therefore cannot be male
• Neck webbing and widely spaced nipples
Intelligence and lifespan is normal
47, XXY
Klinefelter syndrome
• Tall stature, long limbs
• Male but infertile, small testes, about 50% gynaecomastia
Mild learning difficulties
types of structural abnormalities
- Balanced or unbalanced rearrangements
- Translocations (Reciprocal/Robertsonian)
- Deletions
- Insertions
- Inversions
balanced translocation
Non-homologous chromosomes
correct sequences of DNA are present in the correct copy number throughout the entire genome
Reattached in the wrong place but the right amount of DNA is still present in the genome
Generally have little negative effect unless by chance an essential gene was hit during the relocation (very rare)
reciprocal translocation carrier outcomes
formation of two new derivative chromosomes (exchange of the material between 2 chromosomes)
Much more likely to give problems in the next generation where it has occurred in the germ line
Unbalanced genome - missing part of one of the chromosomes but has an extra copy of part of the other chromosome
Depending on the precise nature of this it can give rise to serious defects in the zygote
unbalanced translocation
Balanced translocation can give rise to unbalanced translocation in the next generation
robertsonian translocation
fusion of two acrocentric chromosomes
Translocation can lead to loss of short arms and one large chromosome forms containing all of the coding information from the q arms
acrocentric chromosome
centromere close to one end of the chromosome, p arm is very short (little useful DNA)
robertsonian translocation carrier outcomes
Where gametes inherit the Robertsonian chromosome and one of the normal chromosomes –> large trisomy/ monosomy
Trisomy 14, monosomy 14 and monosomy 21 aren’t compatible with full term pregnancy
Trisomy 21 - duplication of all the info from chromosome 21, this is the causative aetiology of Down’s syndrome
deletions
shorter chromosome arises as genetic material is removed
inversions
balanced rearrangement
piece of DNA is turned around
pericentric inversion
involving the centromere
genetic mutations
• Germline or somatic • Gene disruption /disease-associated (cancer) • Polymorphism ○ No phenotypic effect Frequency >1%
types of genetic mutations
• Non-coding • Coding ○ Silent ○ Missense ○ Nonsense ○ Frameshift
silent mutations
– synonymous e.g. CGA (Arg) to CGC (Arg) - no change in the phenotype
missense mutations
e.g. CGA (Arg) to GGA (Gly) - depending on the biochemistry this may/may not have a serious effect
nonsense mutations
e.g. CGA (Arg) to TGA (Stop) - shortens the protein, will probably get degraded
frameshift mutations
deletion / insertion e.g. CGA (Arg) to CCGA (Pro, then out-of-frame) - changes the reading frame of the rest of the sequence
mutation nomenclature: Cys64Arg
1294del40 - 1294
1298A>G
cysteine at position 64 changed to arginine
-1294 position, deletion of 40 nucleotides at this position
at position 1298 in the nucleotide sequence, A changed to G
detecting mutations
• Polymerase chain reaction (PCR)
• Gel electrophoresis
• Restriction fragment length polymorphism (RFLP) analysis
• Amplification refractory mutation system (ARMS)
DNA sequencing
PCR
amplification of a small amount of DNA
what is needed for PCR
• Sequence information • Oligonucleotide primer • DNA • Nucleotides DNA polymerase
gel electrophoresis
- Separate DNA fragments by size
- Apply an electric field
- DNA is negatively charged
- Separate through agarose gel matrix
- Visualise DNA fragments
advantages of gel electrophoresis
• Speed
• Ease of use
• Sensitive
Robust
PCR applications
DNA cloning, DNA sequencing, In vitro mutagenesis, Gene identification, Gene expression studies, Forensic medicine, Typing genetic markers, Detection of mutations
ARMS
Amplification refractory mutation system
simple method for detecting any mutation involving single base changes or small deletions
pros/cons of ARMS
• Cheap • Labelling not required • Electrophoresis required • Primer design critical Need sequence information Limited amplification size
restriction endonuclease
• Enzymes from bacterial cells • Protective mechanism • Degrade DNA of invading viruses • Recognise specific DNA sequences • Usually 4-8 bp Always cut DNA at the same site
RFLP analysis
involves cutting a particular region of DNA with known variability, with restriction enzymes, then separating the DNA fragments by agarose gel electrophoresis and determining the number of fragments and relative sizes.
restriction digest assay
procedure used in molecular biology to prepare DNA for analysis or other processing
uses restriction endonucleases to cleave DNA at particular sequences
advantages/disadvantages of RFLP
• Simple • Cheap • Non-radioactive • Requires gel electrophoresis Not always feasible
DNA sequencing
• Chain termination method (Sanger)
Use of dideoxynucleotides
pros/cons of DNA sequencing
• Gold standard for mutation detection • Automation and high throughput • Expensive equipment • Poor quality sequence read ○ First part of sequence (15 to 40 bases) ○ Deterioration after 700-900 bases • Next generation sequencing 18 billion bp in 4 days (about 6 human genomes)
