We're going to start a second series of podcasts on cytogenetic. So, we consider
these podcasts part two of our series on cytogenetics. And there are generally going to
deal with the chromosome structure and abnormalities. So, the learning objectives
for this series of podcasts: you should be able to explain chromosome
nomenclature, what chromosome banding is and how karyotypes
are obtained, you should be able to explain linkage analysis in relation to
recombination, you should be able to explain implications of mitotic
nondisjunction with respect to aneuploid genotypes, and finally, we hope you can
appreciate how chromosome structure is altered by balanced or unbalanced
chromosomal rearrangements. Let me just set the stage for this next series of
podcasts. If we consider chromosomal abnormalities,
they likely occur in one in a hundred and fifty live births. Chromosomal
abnormalities contribute significantly to genetic diseases. So, for example, there
are leading cause of intellectual disability. Chromosomal abnormalities
contribute significantly to cancer, especially, if the abnormalities occur in
somatic cells. I highlight a review article that you might want to look at
from the New England Journal of Medicine. Although it's a little bit old now, it is
a good article to talk about chromosomal abnormalities and somatic cells as
related to cancer. Chromosomal abnormalities account for a significant
percentage of pregnancy loss by spontaneous abortions and indeed 50% of
first trimester pregnancy losses likely occur because of a chromosomal
abnormality. In this case, these losses occur very early, oftentimes a woman
doesn't even know she's pregnant. Twenty percent of second trimester spontaneous
abortions are likely a result of chromosomal anomalies. We don't think
about this, but third of pregnancies are lost spontaneously. Now, I'd like to spend
the remaining time in this podcast dealing with linkage. Before I do that,
let's just highlight things that I think you appreciate
from classical genetics. So, we talk about a gene as the fundamental unit of
inheritance and usually corresponds to the segment of DNA coding for a single
protein and a genome is an organisms set of genes. The locus of a gene is the site
of the gene in the genome and alleles would be alternative forms of a gene and
we know that we have usually two alleles for each gene because we have homologous
pairs of chromosomes. The genotype is the specific set of alleles forming the
genome of an individual and the phenotype is the visible character of
the individual. Now, we can talk about the wild type form would be the normally
occurring type versus the mutant form would be different from the wild type
because of a genetic change, usually a mutation. Now, in this example with the
fishes, allele A is dominant relative to allele a, which is considered
recessive. So, in this example, the phenotype of the heterozygote Aa
is the same as that of homozygous dominant AA. In cases
where it would be different from both the two alleles are said to be
co-dominant. So, for example, the ABO and MN blood groups in humans are a result
of co-dominant alleles. This is something that I think you appreciate is well. Here
was showing a chromosome at the beginning of the cell cycle in the G1
phase and this long single bar represents one long double helix of DNA.
So, here's the centromere, the short arm, and the long arm. This part of the
diagram shows a chromosome at the end of the cell cycle in metaphase. It's
duplicated and condensed consisting of two identical sister chromatids each one
containing a DNA double helix and they're joined at the centromere. A
normal diploid chromosome set is seen in metaphase spread. It would be prepared by
bursting open a cell and at metaphase and staining the scouted chromosomes.
So, in the example shown here, there are three pairs of autosomes like this
maternal and paternal chromosome one, maternal and paternal chromosome two,
maternal and paternal chromosome three, and here a pair of sex chromosomes. If
this was from a human cell there'd be 22 pairs of autosomes and a single pair of
sex chromosomes. And, this is not something you need to worry about for
the course, but I just like to put it in perspective, our closest primate relative,
the chimpanzees, have 23 pairs of autosomes; we have 22 pairs of autosomes.
Let's talk a little bit more specifically about linkage. This is a
simple-minded diagram. It's looking at meiosis and genetic recombination. Here
it's showing two loci on a maternal chromosome, two loci on the paternal
chromosome. We come to appreciate that the greater the distance between two loci
on a single chromosome, the greater the chance that they will be separated
by crossing over. And if two genes are assorted in a certain percentage of
gametes, they're said to be separated on a chromosome by a certain genetic map
distance or map units. We appreciate that linked genes are not separated by
crossing over. Now, for human disease, genetic analysis doesn't usually look at
genes but it usually looks at related markers on chromosomes so called
single nucleotide polymorphisms. In the example here, these loci AB and ab.
You can have a genotype Ab and then a
genotype aB. This would be a site where these have crossed over. So,
you'd have a genotype Ab, a genotype aB. The concepts on
linkage are becoming more important and understanding predictive value of
determining linkage between genetic markers and disease
causing gene. I'm going to review some things for you here. They're not going to
necessarily be testable in our course, but it's important to have this as
background information to understand linkage. Linkage is considered the
closeness of two or more loci on a chromosome that sort together during
meiosis one. Now, a loci would be a location on a chromosome of a
disease-causing gene and some non related marker. Linkage would exist
because crossover or recombination rarely occurs between loci that are
close together. If crossover occurs between two loci,
a recombinant chromosome is formed. And we can talk about a recombination
fraction as the number of recombinant chromosomes divided by the total number
of offspring. This seems kind of odd to put it in this these terms, but consider
a family has eight children. Chromosome analysis reveals that seven children had
parental type chromosomes, that is a crossover did not occur between two loci
under consideration, and one child has a recombinant chromosome. That means
a crossover did occur. So, one could say in this hypothetical example, the
recombination fraction is one eighth or 13% or about really
12.5%. A centimorgan defines the genetic distance between two loci.
One centimorgan is defined as the distance between two loci that
produce a recombination fraction of 0.01 or recombination
percentage of 1%. One centimorgan is the length of 1 million
base pairs of DNA on a chromosome. Just for convenience, there are a combination
fraction is sometimes given the term theta. Remember, 1 centimorgan is 1
million base pairs of DNA. That gives a recombination fraction of 0.01
or a recombination percentage of 1%. If we talk about high linkage, we say two
loci are 0.01 centimorgans apart. That would mean the recombination fraction is
0.001 with a recombination percentage of 0.1%. This means there is a 0.1%
chance or one chance in a thousand of recombination. That really
means if a thousand gametes are produced during gametogenesis one gamete will
contain a recombinant chromosome. Remember, if genes are linked recombination
is not likely. Intermediate linkage is considered two loci that are 1
centimorgan apart. That would be equivalent to a recombination fraction
of 0.01 or a recombination percentage of 1%. That would mean that if
a thousand gametes are produced, 10 will contain a recombinant chromosome. If
linkage is low, would say two loci are 50 centimorgans apart. The
recombination fraction would be 0.5, the recombination percentage would be 50%. If
a thousand gametes are produced 500 will contain a recombinant chromosome, that
would be a 50% chance that recombination occurs. Notice that the lower the linkage,
the greater the likelihood of recombination. What is the likelihood the
two loci are linked at a given recombination frequency? Population
geneticists sometimes calculate what's called a LOD score or a LOD of the odds
score. Again, this is not testable for our course,
but this LOD scores is LOD to the base of 10. That the probability the two loci
linked divided by the probability the two loci not linked. If a LOD score is
greater than or equal to 3, that's evidence of linkage between two loci,
if the LOD score is less than or equal to -2,
that's evidence against linkage between two loci. So, a LOD score of
+3 would be a thousand to one that the loci are linked. A LOD score of
-2 would be a hundred to one that the low sigh are not linked. Here's a
hypothetical question for you: the recombination percentage of two low sigh
is 66%, which are the following explanations best fits with
this observation? The two loci are far apart and there is low linkage between
them, the two loci are close together and there is high linkage between them, the
recombination fraction between the loci is low, or the LOD score is high. The
two loci are far apart and there is low linkage between them. That would be
the best choice. With a 66% recombination fraction, these loci are at
least 66 centimorgans or 66 million base pairs apart. Remember, 1 centimorgan
equals a million base pairs, so the recombination fraction is 0.01
or a recombination percentage of 1%. The two loci are close together
and there is high linkage between them. 50% recombination is considered
low linkage. We say here the 3 combination percentage is 66%. The
recombination fraction between the loci is low. If the recombination fraction was
low, these low sighs should be very close together.
Remember, 0.1 centimorgan is about a hundred thousand base pairs. The
recombination fraction would have been 0.001 or 0.1%
recombination. If the loci were absolutely linked, there of course would
be no recombination. So, the clue here, we're saying the recombination
percentage 66% would suggest the loci are far apart. The LOD
score should be low, these loci should not be linked with a 66%
recombination. Now, so the question is this a real
example? It's an interesting point. How many base pairs are there on human
chromosomes? The human genome is something like 3 billion base pairs of
DNA. Chromosome 1, the largest chromosome, is about 250 million base pairs. The Y
chromosome is fairly small, 59 million base pairs. Chromosome 2, the smallest,
has 48 million base pairs. So, here, in this example, 66 million base pairs apart,
so this is not a small chromosome obviously. It could be real if you're
talking about say human chromosome 1, but it's certainly not a real example if
you're talking about some of the smaller human chromosomes. So, linkage analysis,
what does it really mean? If there's absolute linkage, there's zero
centimorgans separating two loci, the recombination fraction theta is zero, there's
0% recombination. So, in meiosis one, as you get synapses occurring you're
going to have one of each of the types of gametes formed: AA, ab,
AB, and another aB, no recombination. If
there's low linkage, the loci are say 50 centimorgans apart, the recombination
fraction theta is 0.5, the recombination percentage is 50% . During synapses at
meiosis one, you'll have crossing over. For a gamete you should have either a AB, a
Ab. Reflecting recombination aB.
Reflecting recombination or ab. So, high linkage means low
recombination. The LOD score be greater than or equal to 3. Low linkage means
high recombination. You have a LOD score of less than -2.