BLGY1232 Locating genes underlying human phenotypes
If the biochemical basis of the phenotype is known
This usually means we know which enzyme is defective and we know something
about the polypeptide sequence of the enzyme
The protein sequence can be used in a genome database search to identify the
gene sequence in the human genome allowing us to PCR-amplify the dene for
affect and unaffected individuals and identify any mutation in the affected
individuals and their families to see if they are also at risk
Haemoglobin in haemoglobinopathies, Hexosaminidase A in Tay-Sachs disease,
Apolipoprotein E in typeIII hyperlipidaemia, Cystic fibrosis etc.
If we don’t know the biochemical basis of a condition
We have to locate where the gene is in the genome, identify the correct coding
sequence and determine the nature of the mutation and its effect on the protein
encoded
2 statistical approaches can be used; analysis based on genetic linkage or
analysis based on allelic association both rely on genetic variation
(polymorphisms) between individuals
To locate the gene, we need to know where it lies in the genome in relation to
other known sequences that differ between individuals - We need to analyse
how it relates to other polymorphisms and the key point is that the variant
sequence should be maintained within a proportion of the population: it’s not
just a one-off mutation
Polymorphism = The existence of two or more variants (alleles, phenotypes, DNA
sequence variants, chromosome structural variations) at significant frequencies
in the population
Autozygosity mapping
Pedigree analysis in families with a high incidence of consanguineous marriage
Where parents are related, there is a higher frequency of autosomal recessive
inherited disorders
We can try and determine the genotypes of affected individuals and their family
members, to identify the genetic interval within which the disease gene lies; this
, region will be homozygous in affected and heterozygous in unaffected family
members
Genetic polymorphism
The existence of 2 or more variants (alleles, phenotypes, DNA sequence variants,
chromosome structural variations) at significant frequencies in the population
Any genetic variant allele that exists at or above a frequency of 1% of the
population
Any non-pathogenic variation
Polymorphisms can be phenotypic (e.g. different blood groups) – but there are
not very many easily identified phenotypic polymorphisms whose exact genetic
location is known
DNA sequence variations are much more useful, because
(1) There are very many throughout the genome
(2) They can be identified by molecular analysis
(3) They are inherited in a codominant manner and we can identify homo- and
hetero-zygotes
RFLPs - restriction fragment length polymorphisms, usually resulting from
mutations that occurred in RE sites; mutation of a restriction enzyme recognition
site results in loss of a cut site
RFLPs are detectable by Southern blotting This is a laborious process, and so is
not used very often for polymorphism analysis using Southern blotting we can
detect different forms (alleles) of individual genes in individual members of
populations
If the biochemical basis of the phenotype is known
This usually means we know which enzyme is defective and we know something
about the polypeptide sequence of the enzyme
The protein sequence can be used in a genome database search to identify the
gene sequence in the human genome allowing us to PCR-amplify the dene for
affect and unaffected individuals and identify any mutation in the affected
individuals and their families to see if they are also at risk
Haemoglobin in haemoglobinopathies, Hexosaminidase A in Tay-Sachs disease,
Apolipoprotein E in typeIII hyperlipidaemia, Cystic fibrosis etc.
If we don’t know the biochemical basis of a condition
We have to locate where the gene is in the genome, identify the correct coding
sequence and determine the nature of the mutation and its effect on the protein
encoded
2 statistical approaches can be used; analysis based on genetic linkage or
analysis based on allelic association both rely on genetic variation
(polymorphisms) between individuals
To locate the gene, we need to know where it lies in the genome in relation to
other known sequences that differ between individuals - We need to analyse
how it relates to other polymorphisms and the key point is that the variant
sequence should be maintained within a proportion of the population: it’s not
just a one-off mutation
Polymorphism = The existence of two or more variants (alleles, phenotypes, DNA
sequence variants, chromosome structural variations) at significant frequencies
in the population
Autozygosity mapping
Pedigree analysis in families with a high incidence of consanguineous marriage
Where parents are related, there is a higher frequency of autosomal recessive
inherited disorders
We can try and determine the genotypes of affected individuals and their family
members, to identify the genetic interval within which the disease gene lies; this
, region will be homozygous in affected and heterozygous in unaffected family
members
Genetic polymorphism
The existence of 2 or more variants (alleles, phenotypes, DNA sequence variants,
chromosome structural variations) at significant frequencies in the population
Any genetic variant allele that exists at or above a frequency of 1% of the
population
Any non-pathogenic variation
Polymorphisms can be phenotypic (e.g. different blood groups) – but there are
not very many easily identified phenotypic polymorphisms whose exact genetic
location is known
DNA sequence variations are much more useful, because
(1) There are very many throughout the genome
(2) They can be identified by molecular analysis
(3) They are inherited in a codominant manner and we can identify homo- and
hetero-zygotes
RFLPs - restriction fragment length polymorphisms, usually resulting from
mutations that occurred in RE sites; mutation of a restriction enzyme recognition
site results in loss of a cut site
RFLPs are detectable by Southern blotting This is a laborious process, and so is
not used very often for polymorphism analysis using Southern blotting we can
detect different forms (alleles) of individual genes in individual members of
populations
