Genetics in Medicine 3 Flashcards
What is the molecular difference between deoxyribose and ribose sugar and what consequences does this have for the structure?
Ribose sugar has a hydroxyl group on C2 which is missing in deoxyribose
This hydroxyl group gives potential for nucleophilic attack on nearby phosphodiester bonds meaning that RNA is transient (much more unstable) but DNA is much more stable
What direction is DNA written in and DNA and RNA synthesised?
written in 5’ to 3’ direction
When DNA sequence is written in shorthand the sequence of which strand in which direction is the one written out?
The sense strand in the 5 to 3 direction
In which stage of the cell cycle does DNA replication take place?
S phase
Before mitosis
In G1 phase how much DNA do you have?
One paternal chromosome
One maternal chromosome
2n
In G2 phase (after S phase) how much DNA do you have?
One paternal chromosome and one maternal chromosome each made up of 2 sister chromatids
4n
How many strands of DNA is one individual chromosome made up of?
One strand of DNA
How many Mb of DNA do we have per haploid genome?
3000Mb per haploid genome
Which is the largest chromosome?
Chromosome 1
What percentage of DNA is non coding?
> 90%
How many coding genes do we have approxiamtely?
20,000
The average gene size is 50-100kb but the average mRNA is ~ 2kb, why is this?
Because of splicing, introns in the gene are removed leaving just exons
What are the 2 main classes of sequences in the human genome?
Single copy sequences (non repetitive)
Repetitive sequences
What are the 2 types of repetitive sequences on the human genome?
1) interspersed repeats
2) Satellite DNA
Alu repeats are an example of what kind of repeats?
Interspersed repeats
What is meant by satellite DNA?
Large blocks of repetitive sequences - heterochromatin
What is the basic definition of a gene?
Functional units of DNA
What is meant by a gene being expressed?
All genes are transcribed at some point - ie copied into RNA
Some of those are then translated - turning the RNA into a protein, but not all as some are functional as mRNA eg. tRNA or rRNA - have short and long coding RNAs
What are the 3 main component parts of a gene?
1) Exons
2) Introns
3) Regulatory sequences eg. promoters, enhancers, locus control regions
By definition where does transcription begin and where does translation begin?
Transcription starts at the beginning of exon 1
Translation begins at the initiator site for translation
Dont get them mixed up
What is alternative splicing and what can it lead to?
Some exons can be removed during splicing leaving 2 or more possible recombinations for spliced mRNA
This leads to much greater genetic variation from just those 20,000 genes - you can end up with 2 different possible proteins from one gene
What is mutually exclusive exon choice in alternative splicing?
May be a piece of DNA with 2 exons that are identical and sometimes one may be incorporated and the other excluded and visa versa
What is meant by the term gene families?
When you look at how the genome has evolved it has evolved alot through duplication and divergence of ancestory genes - you thus end up with gene families of structurally related genes - these can end up close together on a genome or widely dispersed
What happens in the evolution of genes in terms of DNA duplication and divergence?
1) Start off with an ancestral gene
2) As a result of inaccuracies end up with gene duplication - these are initially tandomly arranged
3) As a result of further mutations you get divergence of these duplications
4) They may then aquire different functions
What are pseudogenes and why can they create problems clinically?
Genes that were once useful but as we’ve progressed through evolution are no longer useful so have been turned off
These can be very similar to functional genes so can interfere with medical diagnosis
What are processed genes and how do they form?
Intron-less copies of other genes which are usually remote from the parent gene
Reverse transcription of spliced mRNA and reintegration into the genome
Do processed genes remain functional?
They occasionally remain functional (eg. PGK2 testis specific) but most are non functional
Roughly how many coding, short non-coding, long non-coding and pseudogenes do we have?
Coding - ~20,000
Short non coding - ~9000
Long non-coding - ~13,000
Pseudogenes - ~14,000
What is meant by interspersed repeats?
Those scattered around the genome
Where is satellite DNA found in chromosomes?
Large blocks are found in all chromosomes around the centromere
Also have heterochromatic chromosomal regions
Above simply refers to the fact that satellite DNA stains differently so is called heterochromatin (different to other chromatin) - heterochromatic regions are those made up of satellite DNA
What is alphoid DNA?
Type of satellite DNA found at centromeres
Alphoid DNA is required for the assembly of the centromere
What is significant about the sizes of blocks of satellite DNA in different peoples genome?
Its polymorphic
What is a polymorphism?
Variation in the genome present in atleast 1% of the population
How many bp is a unit of alphoid DNA made up of?
171 bp
How can alphoid DNA be useful in studying chromosomes?
The repetitive sequence shows chromosome-specific sequence variation
Can make probes for individual chromosome identification
How much of the genome is made up from Alu repeats?
5%
How have the Alu repeats become dispersed within the genome and why are Alu repeats clinically significant?
Dispersed by retrotransposition
They have a large role in the generation of molecular pathology
During what process do most mutations occur?
During gametogenesis
At what point in gametogenesis do errors tend to lead to large insertions or deletions tend to occur?
During crossing over and exchange of material between the maternal and paternal homologues
What is a chiasm?
Point at which the maternal and paternal homologues cross over during meiosis 1 of gametogenesis
Describe the process by which interspersed repeats can lead to large deletions or insertions during the process of crossing over?
1) When homologues align they undergo a recognition process which is sequence dependent
2) Problems arise when the sequence specificity of the alignment goes wrong
3) May have 2 Alu elements sitting in adjacent introns, can get chiasm forming and crossing over between the 2 non corresponding regions of Alu
4) You then end up with a non equal cross over and imbalanced products - one has a large duplication and one has a large deletion
What happens in meiosis if crossing over doesnt occur?
Crossing over is an obligatory process - if it doesnt occur then meiosis fails
In a large deletion what is the difference between an in-frame and an out of frame deletion?
Depends on the size of the exon deleted
If the number of base pairs in the exon is a multiple of 3 you will lose however many amino acids are coded for by that particular exon but there is not frame shift - in frame deletion
If the number of base pairs in the exon deleted is not a multiple of 3 then you lose the amino acids coded for in that exon plus the reading frame is shifted for all the following exons so you alter all amino acids coded for by them
Why is an out of frame deletion often more deleterious than an inframe deletion?
Because of the reading frame shift not only are you likely to alter many more amino acids with an out of frame deletion and furthermore by chance are very likely to create a stop codon leading to a truncated problem - greater loss of function
Duchenne muscular dystrophy is caused by what kind of mutation?
Large deletion
Charcot-Marie-Tooth disease is a nerve conduction disorder that results from what kind of mutation?
Large insertion
Why can large deletions or insertions often be missed by PCR methods?
As they dont change the actual sequence of nucleotides except at the breakage point
PCR and arrays that detect the loss or absence of a sequence dont pick up that there is one too many or one too few
Give an example of a gross rearrangement?
Haemophilia A mutation
When do most mutations occur?
Gametogenesis
What is Haemophilia A - what gene/chromosome is involved?
Deficiency of coagulation - deficiency in Factor 8
Gene for Factor 8 is on X chromosome - condition is X linked
What happens in the Haemophilia A mutation?
Factor 8 has a repetitive sequence which is also present further down in the chromosome in the opposite direction
When the gene is copied you alignment of the 2 repetitive sequences and you get inversion of the repeat segment into the middle of factor 8 gene - so factor 8 gene is cut in half
This is intrachromosome recombination
What is a point mutation?
Substitution of one nucleotide
What is a silent point mutations?
Substitution of a nucleotide that results in no alteration to the amino acid sequence
What is a misense point mutation, what is the difference between conservative and non conservative?
Substitution of a nucleotide results in a change to the amino acid sequence
Conservative - eg. change for one polar amino acid for another polar amino acid
Non-conservative - change from an acidic amino acid to a basic amino acid
What kind of substitution makes up one 3rd of point mutations?
CG -> TG
Why does CG->TG make up 1/3 of mutations?
Site of substitution is most commonly CpG
As this is where DNA gets methylated
Methylcytosine by simple deamination can be converted to Thymine
Its difficult for mutation checking systems to know whether the T is wrong
What is a non sense point mutation?
Where the substitution of a nucleotide results in the production of a mis placed stop codon
A truncated protein is produced
This can be exploited for mutation detection - protein truncation test
What is a frameshift mutation?
Deletion or insertion of one nucleotide
How can you get mutations outside of exons that cause disease?
Because they alter splicing
The first and last codes of introns are critical to determine where splicing occurs
If these are altered on the intron, incorrect splicing can occur and you may get large insertions into mRNA
When describing mutations a reference sequence (or what sequence should be) is always given, what 3 kinds can be used?
1) Genomic DNA
2) cDNA - made from mRNA
3) Protein
What would p.Ala2Val mean?
Protein is the reference sequence
The second amino acid in the protein has been changed from Alanine to Valine
What would c.658delG mean?
cDNA is the reference sequence
A guanine nucleotide in the 658th position has been deleted
What would c.[44C>T] ; [826G>A] mean?
In recessive conditions is showing the maternal and paternal allele, separated by a semi colon
This shows cDNA is the reference sequence in one allele the 44th nucleotide has changed from a cytosine to a thymine and in the other allele the 826th nucleotide has changed from a guanine to an alanine nucleotide
Are mutations that result in loss of function normally recessive or dominant and why?
Recessive
Can cope with 50% loss of function in that gene
Are mutations that result in gain or alteration of function normally recessive or dominant and why?
Dominant
It causes a receptor or protein or enzyme to be expressed more than it would normally be
What is meant by compound heterozygotes for recessive disorders and why can this be challenging for clinically?
Have 2 different recessive alleles for a disease- are affected as have no functioning gene
This can be challenging for mutation testing as have to have knowledge of many different mutations
Give 2 examples of recessive disorders?
1) CF
2) Beta Thalassemia
Do dominant conditions have a greater or lesser mutational spectrum that recessive conditions?
Lesser - less chance of mutations that will result in gain of function as in the majority of dominant conditions than mutations that will result in loss of function as in the majority of recessive conditions
Give one example of a dominant condition caused by a mutation which results in gain of function?
Achondroplasia
What are trinucleotide repeat expansions?
Class of mutations that result from a string of repeats of 3 nucleotides - you get an incorrect expanded number of the repeats causing disease
What are the 2 main types of trinucleotide expansions and name 2 diseases that each causes?
1) Polyglutamine repeats (CAG) - get many CAG repeats leading to along string of glutamine in protein, alot of diseases resulting from these are neurodegenerative
- Huntington’s disease
- Spinocerebellar ataxias
2) Large non-coding repeat expansions - increased number of repeats in non coding regions but this alters regulations of the gene
- Fragile X syndrome - transcriptional silencing
- Myotonic dystrophy
Trinucleotide expansions show mutational instability, what is meant by this?
The mutation as it is passed through generations can gradually get further expanded, seeing an increasing number of repeats as it is passed from generation to generation
This is occasional in Huntington’s disease but frequent in Fragile X syndrome
In Fragile X syndrome, what is the difference between an unstable mutation and full mutation?
Defined by the number of trinucleotide repeats
In the process from going from a normal number to full mutation there is a series of steps and normal becomes an unstable mutation with more than a normal number of repeats but not quite as many as a full mutation before it becomes a full mutation
