Molecular basis of diseases
3rd year biology
Lecture 1: Molecular genetics: introduction to human genetics
Genomic variation:
- Genomic terminology
- Genome is variable
Intellectual disability:
- ID is a social disease
- ID is heterogenous
Genomic terminology:
- Variation: any deviation from the reference genome
- Polymorphism: variation ≥ 1% of the alleles in the population
- We all have 2 alleles, so in a test with 50 people you have 100 alleles
- Carrier frequency in this group can be different from allele frequency (1 in 50 carrier and 1 in
100 alleles)
- Mutation: variaton ≤ 1% of the alleles in the population
- Pathogenic: disease-causing mutation or polymorphism
- CNV: deletion or duplication ≥1 kb
Reference genome= are 6 persons that are sequenced, we always map back to these persons
- Contains 3 billion basepairs (1 allele),
stored in genome browsers
- 6 billion bp = 2 alleles
Genome variation:
- Single nucleotide variant (SNV): basepair
change (or SNP)
- Insertion or deletion (indel): ≤ 10 bp
- Repeat expansion: 2 to > 6000 bp
- Copy number variant (CNV): deletion or
duplication > 1000 bp
- Structural chromosomal abnormalities:
translocation, inversions, etc.
- Aneuploidy: one chromosome extra
Splice site changes: splice site can possibly not be
recognized if changes happen at the splice site.
Acceptor splice site: Always an A and then a G
before the exon starts.
Donor splice site: Always a G and then a T after the exon
1
, Molecular basis of diseases
3rd year biology
Intellectual disability (ID):
- Intelligence quotient (IQ) < 70
- Limitations in adaptive functioning
- Present < 18 years
ID classification
Class IQ Education
Profound <20 Institutionalised -> no education
Severe 20-34 Special education
Moderate 35-49 Special education
Mild 50-69 Normal education, until a certain age and they cannot keep up
Causes of ID:
- Multifactorial: multiple gene packages that have to be changed before you get the disease
- Organic: you can actually see that there is something wrong with the brain
o We now know it is monogenic: single abnormal gene
ID inheritance:
- Often without family members that have ID is it recessive, or de novo, can it be x-linked?
- Heterogeneous
ID is a heterogeneous disorder
Allelic heterogeneity:
- Same/different phenotype from an identical gene
- Same gene
- Different mutation
2
, Molecular basis of diseases
3rd year biology
Locus heterogeneity:
- Same phenotype
- Different genes
Genome-wide detection of variation:
- Karyotyping - CNVs
- Chromosomal microarray - CNVs
- Exome sequencing – SNVs, Indels, CNVs
- Genome sequencing - SNVs, Indels, CNVs
Lecture 2: intellectual disability: arrayCGH
Cytogenetic SNP array analysis
Cytogenetic diagnostics for ID patients:
Chromosomal anomalies are a well-known cause of ID
Multiple genes are often affected which will affect brain development
3
, Molecular basis of diseases
3rd year biology
Before 2009, they used karyotyping: looking at
chromosomes through microscopes. Prepare
chromosomes in metaphase. You can then see if
chromosomes are intact or not. Are pieces missing or
added?
Karyotyping: limited resolution:
- You only see abnormalities when they involve
millions of nucleotides
o Resolution: ~500 bands
o ~1 band per 6 million nucleotides
- (Much smaller aberrations can cause aberrant
phenotype as well)
- It is time-consuming, difficult to automate,
interpretation is subjective
Analysing chromosomes indirectly to find very small
aberrations (sub microscopic aberrations). Micro syndromes.
1. You have to recognize the phenotype, e.g. Down
syndrome.
2. Analyse the region of interest in the genome, e.g. can
you spot Trisomy 21?
3. Targeted analysis by for example FISH
Fish methodology: Fluoresence In Situ Hybridization
1. Chromosomes on a microscope slide
2. Make DNA single stranded by heating the DNA
3. Add a probe directed against the chromosome/region
of interest
4. Look through the microscope if you can see a
signal at that position
In Wolf Hirschhorn syndrome (deletion 4p)
- Recognize patient by phenotype ask for FISH
- Probe against 4p
- Green indicates centromere of chromosome 4
- Red indicates 4p region missing on 1
chromosome
Genome diagnostics for ID with or without multiple congenital anomalies (MCA)
4
3rd year biology
Lecture 1: Molecular genetics: introduction to human genetics
Genomic variation:
- Genomic terminology
- Genome is variable
Intellectual disability:
- ID is a social disease
- ID is heterogenous
Genomic terminology:
- Variation: any deviation from the reference genome
- Polymorphism: variation ≥ 1% of the alleles in the population
- We all have 2 alleles, so in a test with 50 people you have 100 alleles
- Carrier frequency in this group can be different from allele frequency (1 in 50 carrier and 1 in
100 alleles)
- Mutation: variaton ≤ 1% of the alleles in the population
- Pathogenic: disease-causing mutation or polymorphism
- CNV: deletion or duplication ≥1 kb
Reference genome= are 6 persons that are sequenced, we always map back to these persons
- Contains 3 billion basepairs (1 allele),
stored in genome browsers
- 6 billion bp = 2 alleles
Genome variation:
- Single nucleotide variant (SNV): basepair
change (or SNP)
- Insertion or deletion (indel): ≤ 10 bp
- Repeat expansion: 2 to > 6000 bp
- Copy number variant (CNV): deletion or
duplication > 1000 bp
- Structural chromosomal abnormalities:
translocation, inversions, etc.
- Aneuploidy: one chromosome extra
Splice site changes: splice site can possibly not be
recognized if changes happen at the splice site.
Acceptor splice site: Always an A and then a G
before the exon starts.
Donor splice site: Always a G and then a T after the exon
1
, Molecular basis of diseases
3rd year biology
Intellectual disability (ID):
- Intelligence quotient (IQ) < 70
- Limitations in adaptive functioning
- Present < 18 years
ID classification
Class IQ Education
Profound <20 Institutionalised -> no education
Severe 20-34 Special education
Moderate 35-49 Special education
Mild 50-69 Normal education, until a certain age and they cannot keep up
Causes of ID:
- Multifactorial: multiple gene packages that have to be changed before you get the disease
- Organic: you can actually see that there is something wrong with the brain
o We now know it is monogenic: single abnormal gene
ID inheritance:
- Often without family members that have ID is it recessive, or de novo, can it be x-linked?
- Heterogeneous
ID is a heterogeneous disorder
Allelic heterogeneity:
- Same/different phenotype from an identical gene
- Same gene
- Different mutation
2
, Molecular basis of diseases
3rd year biology
Locus heterogeneity:
- Same phenotype
- Different genes
Genome-wide detection of variation:
- Karyotyping - CNVs
- Chromosomal microarray - CNVs
- Exome sequencing – SNVs, Indels, CNVs
- Genome sequencing - SNVs, Indels, CNVs
Lecture 2: intellectual disability: arrayCGH
Cytogenetic SNP array analysis
Cytogenetic diagnostics for ID patients:
Chromosomal anomalies are a well-known cause of ID
Multiple genes are often affected which will affect brain development
3
, Molecular basis of diseases
3rd year biology
Before 2009, they used karyotyping: looking at
chromosomes through microscopes. Prepare
chromosomes in metaphase. You can then see if
chromosomes are intact or not. Are pieces missing or
added?
Karyotyping: limited resolution:
- You only see abnormalities when they involve
millions of nucleotides
o Resolution: ~500 bands
o ~1 band per 6 million nucleotides
- (Much smaller aberrations can cause aberrant
phenotype as well)
- It is time-consuming, difficult to automate,
interpretation is subjective
Analysing chromosomes indirectly to find very small
aberrations (sub microscopic aberrations). Micro syndromes.
1. You have to recognize the phenotype, e.g. Down
syndrome.
2. Analyse the region of interest in the genome, e.g. can
you spot Trisomy 21?
3. Targeted analysis by for example FISH
Fish methodology: Fluoresence In Situ Hybridization
1. Chromosomes on a microscope slide
2. Make DNA single stranded by heating the DNA
3. Add a probe directed against the chromosome/region
of interest
4. Look through the microscope if you can see a
signal at that position
In Wolf Hirschhorn syndrome (deletion 4p)
- Recognize patient by phenotype ask for FISH
- Probe against 4p
- Green indicates centromere of chromosome 4
- Red indicates 4p region missing on 1
chromosome
Genome diagnostics for ID with or without multiple congenital anomalies (MCA)
4
