Genetics 5 - Haplotypes and Consanguinity Flashcards
learning outcomes
gene
what is the determinant
a biological determinant of a Mendelian character (trait)
a functional unit of DNA is now understood to be that determinant
haplotype (haploid genotype)
set of polymorphisms (set of polymorphic genes = alleles OR set of genetic markers = SNPs) that are grouped tightly together on a single chr and tend to be inherited together through many generations (not separated by crossing over)
can refer to a combination of alleles or to a set of single nucleotide polymorphisms (SNPs)/genetic markers
also known as a DNA/genetic signature
chunk of chr (crossing over)
haplogroup
group of similar haplotypes that share a common ancestor with a SNP mutation
e.g. mtDNA haplogroups (7 daughters of Eve) share common SNPs in mtDNA
white eye colour on fruit fly drosophila
linked to X chr
⇒ genes were linked - carried on specific chromosomes and inherited together
why are some “linked” genes not inherited together
crossing over
first chromosomal linkage map ⇒
genes more closely grouped on chromosomes were separated less frequently by crossing over
closely linked genes (e.g. vermillion eyes and miniature wings were more likely to belong to the same haplotype (chunk)
how to show linkage of a haplotype to a character
examine haplotypes of related individuals with the same traits
if they have a haplotype in common, we can assume that the gene responsible for the shared trait is in that area
UNKNOWN - which part of genome we need to focus on - comparing all the genes is very difficult
human nuclear genome - what do we know
lots of noncoding sequences - 98%
including tandem (head to tail) repeats
interspersed repeats (throughout genome i.e. not head to tail, may be on different chromosomes
define tandem repeat
series of nucleotides directly repeated
length of repeat unit varies
can be used as genetic markers for identification of haplotypes
satellite DNA (type of tandem repeat)
size
use
very large arrays of non-coding tandemly repeating DNA
array of 100 kb - several Mb (this is the total size of repeat units put together)
length of repeat unit > 100 bp
sequences vary between individuals
undefined role in cell cycle - used in DNA fingerprinting
minisatellite DNA (type of tandem repeat)
size
use
medium size
array from 10 bp up to 20kb
length of repeat unit = 10-100 bp
high mutation rate, high diversity in population - VNTR (Variable Number of Tandem Repeats) ⇒ the more it mutates, the more it repeats, the more common it is that the mistake will happen
used for variable tandem repeat analyses
found in subtelomeric region of chr ⇒ protects chr ends from damage
microsatellite DNA (type of tandem repeat)
size
use
simple sequence repeats (SSR) or short tandem repeats (STR)
arrays of less than 100 bp
length of repeat unit typically 2-4bp
used in genetic linkage analysis to locate a gene or a mutation responsible for a given trait or disease
MOST COMMON
3 types of tandem repeats
- satellite DNA
- minisatellite DNA
- microsatellite DNA
where is microsatellite DNA found
how much of genome does it account for
type of sequences
dispersed throughout chr
accounts for about 2% of genome (60 Mb)
microsatellites usually in intergenic sequences or introns
sometimes in coding sequences (exons)
microsatellites in exons
tend to be mutation hot spots
instability of microsatellite DNA
during replication DNA polymerase tends to make errors in copying repeated units - due to their similarity
e.g. may skip over a repeat unit or copy it twice
because replication of repeat units is very error prone microsatellite DNA sequences are highly polymorphic (many different forms)
microsatellite polymorphism
other type of microsatellite polymorphism - SNPs
where does it occur
occurs upstream of a microsatellite
SNP in Man 1 will not be apparent if the PCR products are analysed only by size, only becomes apparent if you sequence
SNP vs microsatellite polymorphism
microsatellite polymorphisms - look at size
SNP - need to know sequence
using microsatellite DNA as a marker for tracking
suppose human genome databases show a microsatellite (trinucleotide repeat CTT) in intron 2 of gene X on chr 1
can use genome database to design complementary oligonucleotide primers upstream (5’) and downstream (3’) of this microsatellite sequence - flanking sequence
functions of microsatellite sequences as markers
design primers
determine appropriate conditions (annealing) for amplification
extract DNA from patient
amplify the interval between the primers
get PCR products
measure size of PCR product
microsatellite sequence - what fills in the gaps
DNA polymerase
based on size of PCR product, what can you determine
how many repeats or what you’re microsatellite no of repeats is
Primers = 20 nucleotides each
In between is the rest
64 - 20 = 44 24 bps
Each of bp is 3 nucleotides
24 divided by 3 = 8
equation for no of repeats
Size of primer + size of amplified microsatellite sequence in between and divide by no of nucleotides in a repeat unit = no of repeats
tracking chromosomes or chunks of chr
using microsatellites and SNPs you can construct a “barcode” for each Chr or chunk of chr (haplotype)
linkage
relationship between loci
specifically genetic phenomenon
linkage analysis
looks at physical chunks of the genome (haplotypes) of related individuals and associates them with given traits
how are trait causing mutations inherited
jointly (linked) with the genetic markers (i.e. microsatellites and SNPs) located in their immediate vicinity on the same chromosomal strand
principle of linkage analysis
if we find a common genetic marker (e.g. microsatellite or SNP), we assume that the gene that causes the disease is somewhere in the same area
how to identify trait-related haplotype (chunk)
perform genetic analysis of polymorphic DNA markers associated with the trait for all family members
can use the dbSNP database to find appropriate markers
see which markers are carried only by members with the trait
high probability (odds) that trait causing gene is linked to this marker (same location on chr)
red hair as a recessive trait
what pattern would Hermione show
things to remember
learning outcomes
consanguinity
being descended from the same ancestor as another person
8.5% of children worldwide - consanguinous parents
endogamy is widely practised in Middle East
clinical definition of consaguinity
matrimony (breeding) between 2 family members who are 2nd cousin or closer
associated with increased risk of autosomal recessive disorders
risks associated with consanguinity
increased risk of genetic disease from 2 → 4%
probability of having a child without a constitutional congenital defect reduced from 98% → 96%
this pedigree is suggestive of what pattern of inheritance
autosomal recessive deafness
(Do the parents carry it? No - recessive
If females have it, it is unlikely to be X linked)
autosomal recessive deafness accounts for
65% of congenital deafness
infection is also important
syndromic vs non-syndromic deafness
syndromic - part of another condition
4 patterns of inheritance of deafness
dominant (DFNA) - 20-25%
recessive (DFNB) - 75-80%
X-linked (DFNX) - 1-2%
matrilineal (mitochondrial) - <1%
autosomal recessive patterns of inheritance are related to
homozygous state for a defective allele
assumption for inherited deafness
likely there is an inherited determinant of deafness in the family - bad gene
mapping is about finding the DNA sequence that corresponds to this gene
common genes for hereditary deafness
single locus - DFNB1 (13q) accounts for a high proportion
GJB2 (connexin 26 protein)(Cx26) - 2 exons ⇒ amplification and sequencing is practical
c. 35delG - common in Europeans
c. del235C - common in Chinese
after looking for common mutations, what is next
(> 100 genes known associated with hereditary deafness - almost impossible)
basic hypothesis
Nasreen has ended up with 2 copies of GGF’s defective chr 1p
if it was simple what should all 3 deaf individuals have
2 practically identical copies of relevant ancestral chr that carries the defective allele or bad gene
however, crossing over between maternal and paternal means that many offspring do not inherit a full intact version
should all be homozygous for a particular chunk (haplotype) of ancestral chr that carries the defective allele
how to identify deafness-related haplotype by linkage analysis
perform genomic analysis of polymorphic DNA markers associated with deafness for all family members
see which markers are carried only by deaf members and never healthy members (segregate with the disorder)
high probability that deafness causing gene is linked to this/these marker(s) - same location on chr
autozygosity mapping
- identify microsatellite sequences associated with deafness loci using dpSNP database
- amplify DNA from all of the loci of all family members
- determine the size of PCR products in each individual - affected individuals should all be homozygous for the same haplotype (set of markers)
microsatellite locus for Nasreen and her 2 deaf uncles
single products suggests that they are all homozygous for a particular chr 2 haplotype with the same no of repeats (13) at that specific micro-satellite locus
may have 2 copies of an ancestral chr 2 from a specific grandparent
MUMTAZ MAY HAVE 2 DIFFERENT PRODUCTS (heterozygous)
- a 99bp product - ancestral chr 1
- maybe an 81 bp product (7 repeats) from the corresponding locus on the other chr 1 homolog
where is the affected deafness allele located
defective allele for inherited deafness
chr 2 haplotype - a chunk of chr on short arm (p) of chr 2
defective allele is in p22-p23 region of chr 2
deafness associated locus there = OTOF (otoferlin) - 2p23.3
9th indentified type of autosomal recessive non-syndromic hearing loss
DFNB9
association is not causation re
linkage of a trait with a gene/DNA sequence associated with the disorder is not always important in the pathogenesis of the condition
deafness associated locus was OTOF - 2p23.3 and not it’s linked microsatellite marker
Coeffecient of relationship (COR) (Sewall Wright)
proportion of alleles that 2 people share by virtue of having 1 or more definable common ancestors
coefficient of inbreeding (COI)
proportion of loci at which the person is expected to be homozygous because of the cansanguinity of the parents
OR
the probability that at any given locus the person receives 2 alleles that are identical by descent
COI = half the coefficient of relationship of the parents
outbred calculation of COR - parents
share 50% of alleles with their children
outbred calculation of COR - full siblings
50%
outbred calculation of COR - grandparents
share 25% with grandchildren
outbred calculation of COR - uncle/aunt
share 25% with niece/nephew
outbred calculation of COR - cousins
12.5%
allele frequency
incidence of a gene variant in a population
number of times the allele of interest is observed in a population/total number of copies of all the alleles at that locus in the population
reflection of genetic diversity
allele frequency does not ⇒
NOT THE SAME as saying 40% of the population have a copy of the allele
e.g.
10% of 100 people are homozygous for allele R1 (20 copies)
20% are heterozygous for allele R1 (20 copies)
allele frequency = 40/200 = 20%
Hardy-Weinberg Equilibrium
assumptions
allele and genotype frequencies in a population will remain constant from generation to generation in the absence of other evolutionary influences
alleles distributed randomly only under certain assumptions including the absence of selection and random mating (a panmictic population)
p2 + 2pq + q2 = 1
p + q = 1
e.g. Hardy-Weinberg Equilibrium
e.g. Hardy-Weinberg Equilibrium
things to remember
