Question | Answer |
What are 3 exogenous mutagens? | X-rays, uv light, O2 radicals |
What is an endogenous mutation causing event? | Replication errors |
nDNA vs mtDNA - Which has lower mutation rate? | nDNA |
nDNA vs mtDNA - Which has more robust DNA repair mechanisms? | nDNA |
nDNA vs mtDNA - Which is susceptible to damage by reactive O2 species? | mtDNA |
Mutations in DNA coding can be grouped into what two classes? | 1. Synonymous (silent) mutations
2. Non-synonymous mutations |
What are synonymous (silent) mutations? | No change in gene product (degenerate code)
Common |
Are synonymous (silent) mutations common/ rare? | Common |
What are non-synonymous mutations? | Change in sequence of gene product |
Are non-synonymous mutations common/ rare? | Rare |
What are the 4 main instabilities in the human genome? | Base substitutions, insertions, deletions, chromosomal abnormalities |
In base substitutions, what are transitions? | Pyramidine-pyramidine or purine-purine |
In base substitutions, what are transversions? | Pyramidine-purine or vice versa |
What are the 3 types of single base substitutions? | Missense (replaced)
Nonsense (replaced with stop codon)
Spice (alters signal from exon-intron splicing) |
What are multiple base substitutions? | Gene conversion-like events |
Insertions - One or more nucleotides is called what? | Frameshift |
Insertions - Triplet repeat expansions lead to what? | Instability |
Insertions - What are substantial repeat insertions called? | Tandem repeats |
Check you understand DNA libraries | (see notes) |
What are the two major methods of identifying disease genes? | Position independent cloning and positional cloning |
Functional coding - 4 main steps
1. Isolate...
2. Determine... (2)
3. Synthesize...for...
4. Screen DNA library to... | 1. Isolate protein product of a disease gene
2. Determine a.a. sequence of protein and therefore cDNA sequence
3. Synthesize oligonucleotide probes for cDNA
4. Screen cDNA library to recover sequence |
Position-independent DNA sequence knowledge - Requires knowledge of what? e.g. | Disease pathology e.g. Disorders which show 'anticipation', such as those with trinucleotide repeat expansions |
Position-independent DNA sequence knowledge - How is sequence identified? | Southern blotting techniques |
Position-independent DNA sequence knowledge - From the 'anticipation' in the southern blot, what can be identified? | Longer/ shorter expansions |
Positional cloning - What is it? | Isolation of a gene by identifying its precise chromosomal location, by a combination of physical and genetic mapping |
Positional cloning - What are the 5 main steps?
1. Define...
2. Obtain...
3. Identify...
4. Prioritise...
5. Test candidate genes for? | 1. Define the candidate region (linkeage analysis/ chromosomal abnormalities)
2. Obtain clones of all the DNA of the region
3. Identify all the genes in the region
4. Prioritize them for mutation screening |
Linkeage analysis - What are SNPs? | Single nucleotide polymorphism (unique genomic marker) |
Linkeage analysis - Use of SNPSs? | Find an SNP near gene of interest |
Linkeage analysis - What therefore does frequency of the SNP and disease indicate? | The relative proximity of the SNP to the disease causing gene |
Linkeage analysis - How can the exact position of the disease causing gene then be identified? | Method called 'chromosome walking' |
What are cytogenetic abnormalities? e.g. | Chromosomal abnormalities, seen on a karyotype, accompanying a disease e.g. DMD |
DMD - Type of inheritance? | X-linked recessive |
DMD - familial or sporadic? | 2/3 familial, 1/3 sporadic |
What are the three steps in subtraction cloning (use example of DMD)? (3) | 1. Assume large deletion included disease causing (DMD) gene
2. Clone missing DNA from normal x chromosome
3. Fine map region to find DMD gene |