Module A: Cytogenetics Flashcards
What is chromatin?
complexes of chromosomes condensed with histones
Describe the utility of a standard karyotype and how it is obtained.
how it is obtained:
cells are induced to divide, then trapped in metaphase and stained
utility:
allows chromosomes to be visualized under a light microscope
Only certain cells can be induced to divide for a standard karyotype. What are they? (6)
blood lymphocytes
amniotic fluid cells
chorionic villi
skin fibroblasts
testicular biopsies
bone marrow
[BACSTB]
In the absence of genes on a Y chromosome, how will the embryo develop?
embryo will develop into a female
In the context of chromosome staining, what are bands?
predictable patterns of light and dark regions
A typical chromosome spread has how many bands?
550 bands
High resolution chromosome spreads have how many bands?
1500 bands
Define acrocentric chromosome.
centromeres close to one end
Define metacentric chromosome.
centromeres near middle
Define centromere.
constriction points where spindle fibers ttach
Define autosome.
chromosomes that are not sex chromosomes
Define euchromatin.
contains genes being actively transcribed
Define heterochromatin.
condensed and generally inactive
What are homologous chromosomes?
two chromosomes in a pair in diploid organisms
What are satellites?
strings of repetitive DNA (usually rRNA coding) that make up short arms of acrocentric chromosomes
What are telomeres?
strings of repetitive DNA at ends of chromosomes
Human chromosomes are numbered from 1 to 22 in what order?
roughly in size order
Describe how the centromere divides the chromosome.
centromere divides chromosome into short arm (p) and long arm (q)
Describe the band-numbering convention in chromosomes. (4)
bands are numbered extending from centromere (band 10)
first major band = 1, second major band = 2, etc.
subbands of major band given second digit (i.e. third subband of first band = 13)
splitting of subband denoted by number following decimal point
In the standard nomenclature system, how are abnormalities indicated? Give an example by writing out the karyotype nomenclature for a male with Down syndrome. (2)
abnormalities indicated following comma after sex chromosomes
47, XY, +21
In the standard nomenclature system, how are deletions indicated? Give an example by writing out the karyotype nomenclature for a male with a deletion of the short arm of chromosome 5. (2)
indicated with “del”
46, XY, del(5p)
In the standard nomenclature system, how are duplications indicated?
indicated with “dup”
In the standard nomenclature system, how are inversions indicated?
indicated with “inv”
Chromosome abnormalities are relatively common in what groups of people?
couples with infertility
spontaneous abortions/miscarriages
children with multiple congenital anomalies
couples with multiple miscarriages
individuals with cancer
Define aneuploidy.
having abnormal number of chromosomes
Define trisomy.
having three copies of a chromosome
Define triploidy.
having three copies of all the chromosomes
Define monosomy and its clinical relevance. (2)
having only one copy of a chromosome
other than X-monosomy, most monosomies are lethal in humans
Define inversion.
chromosomal anomaly in which part of chromosome is turned around in orientation
Define translocation.
anomaly in which part of one chromosome is attached to another (usually reciprocal)
Define Robertsonian translocation.
attachment of long arms of 2 acrocentric chromosomes at centromere
What is the karyotypic nomenclature for Down syndrome?
47, XX (or XY), +21
What is the karyotypic nomenclature for Kleinfelter syndrome?
47, XXY, +X
What is the karyotypic nomenclature for Turner syndrome?
45, X
Numerical chromosomal abnormalities include (2)
errors of ploidy = multiples of 23 chromosomes other than 2
aneuploidy = extra or missing copies of a single/multiple chromosomes, but not the whole set
Structural chromosomal abnormalities include (5)
reciprocal translocations
Robertsonian translocations
deletions
duplications
inversions
What are copy number variants?
common occurrences of duplications and deletions too small to be seen by standard karyotype analysis
Under what conditions is a chromosome rearrangement called “balanced?”
if there is no missing or extra DNA
Broadly speaking, what are the clinical presentations of someone with a balanced rearrangement?
person should be generally normal, but may be at risk for having children with unbalanced chromosomal rearrangements
In mitosis, what structures behave independently of each other?
in mitosis, the pairs of chromosomes (not the chromatids) behave independently of each other
What happens in meiosis I? (3)
two pairs of chromosomes (each with 2 sister chromatids) pair as bivalents
then homologous regions of the 2 chromosomes pair precisely and cross over
spindle fibers pull bivalents apart, resulting in 2 daughter cells w/ 23 chromosomes (46 chromatids) in each
What is non-disjunction?
failure of chromosomes or chromatids to segregate properly, resulting in aneuploidy
What is mosaicism and how does it develop? (2)
a phenomenon in which some cells have an aberrant genotype, but then other cells within the same individual have a normal genotype
always a post-zygotic event
What is hybridization?
allowing the two strands of DNA to separate, then allowing a matching sequence to bind/anneal to the strand
What is stringency?
how well the sequences must match for a probe to anneal to a DNA strand, dependent on physical and chemical conditions
Describe the properties of short probes. (2)
15-60 nucleotides
requires sufficiently stringent conditions
Describe the properties of longer probes. (2)
~1000 nucleotides
less stringent conditions
Describe the mechanism of Dot blots. (2)
labeled oligonucleotide hybridizes to patient’s DNA
unbound DNA washed off
Dot blots are useful for what purpose?
detecting specific single base changes
Describe the mechanism of Southern blots.
detect the size of a DNA fragment, sometimes produced by cutting DNA w/ restriction endonuclease, that binds to a probe
In what context are Southern blots useful?
used to detect the number of sequence repeats in a gene
FISH stands for
fluorescence in situ hybridization
FISH is used for (2)
determining whether a segment of DNA is present on a chromosome
detecting microdeletions or duplications in patient’s DNA
Describe how FISH is performed. (3)
mitotic chromosome spread placed on slide (just like conventional karyotype)
probe hybridized to slide and excess washed away
fluorescence microscope reveals signal (dot) where probe has hybridized
In what range can FISH detect deletions?
FISH is used to detect deletions in the 40 kb to 1 Mb range
FISH is NOT useful for what kinds of changes? (2)
single base changes
small deletions
Why would interphase FISH be helpful compared to metaphase FISH or conventional karyotypes? (2)
helpful because cells do not need to be induced to divide
study can be done more rapidly than conventional karyotype or metaphase FISH
Describe the process of interphase FISH. (4)
cells bound to microscope slide
probe allowed to hybridize
unbound probe washed away
number of signals (dots) in each nucleus indicates number of DNA sequences
What does CMA stand for?
chromosome microarray
What are the two types of cytogenomic microarray (CMA) analysis?
array comparitive genomic hybridization (aCGH)
in silico methods
What is a common feature of the two types of CMA analysis?
both use thousands of probes bound to a silicon chip in an array
Explain the general basis of comparitive genomic hybridization. (5)
patient + control DNA labeled with different fluorescent probes and mixed
mixture of DNA hybridized to chip with thousands of single copy sequences
washing
if patient DNA has more copies than control DNA, probe will fluoresce with patient color
if patient DNA has fewer copies than control DNA, probe will fluoresce with control color
Typical CMA arrays can detect what size of copy number variants?
20kb or more in size (though more specialized arrays that detect smaller variants do exist)
In CMA analysis, a 1:2 ratio for an autosomal probe indicates
patient has a deletion
In CMA analysis, a 3:2 ratio for an autosomal probe indicates
patient has a duplication
What are single nucleotide polymorphisms (SNPs)?
DNA sequence variations among chromosomes in the population
Describe how SNP-CMA analysis differs from standard aCGH CMA analysis.
patient’s DNA is labeled with a fluorescent probe and washed and all, but this technique does not require control DNA
How is the readout of SNP-CMA analysis interpreted?
intensity of bound fluorescence indicates whether 0, 1, 2 or more sequences from the patient bind to the particular probe
CMA analysis can detect (2)
genetic imbalances
copy number variants
CMA analysis can NOT detect
balanced rearrangements
What is the resolution for conventional karyotyping?
3-5 Mb
What is the resolution for FISH?
100 kb - 1000 kb
What is the resolution for CMA?
5-30 kb
What is the resolution for ASO?
single base differences (but only specific sequence variants)
