SM 17/18 Flashcards
(15 cards)
Why must cytogenetic chromosome analysis must be performed on live, viable cells actively going through mitosis?
Chromosomes are at their most condensed (and therefore most recognizable) size during the metaphase stage of mitosis.
Describe the laboratory methodology for cytogenetic chromosome analysis.
It involves tissue culture (to ensure entrance of cells into mitosis), treatment with a spindle fiber inhibitor (colcemid or colchesine) to block anaphase and a hypotonic solution (low salt) to swell the cells. Cell suspensions are placed on microscope slides and banding using and enzyme to digest protein structures and standard pathology stains (Giemsa and/or Wright).
Name the acrocentric chromosomes.
13, 14, 15, 21, 22
Euploidy vs aneuploidy
Euploidy – a chromosome number which is an exact multiple of a haploid set
Aneuploidy – a number different than 46 but not a whole haploid set different
What would be a normal chromosome name.
46, XX, …
What is the Lyon Hypothesis
X-inactivation
Mediated by the XIST gene which is expressed of the inactive X chromosome and remains in the nucleus as a RNA transcript to coat the inactive X chromosome. . There are a number of genes, however, that escape X-inactivation. Most are located near the telomeres of the chromosomes. The regions are referred to as the “pseudo-autosomal” regions as cells have two expressed copies of these genes just like the genes on the autosomes.
Name the stages of Prophase I
leptotene - chrome begin to condense
zygotene - homologs align (synapse) and held together y synaptonemal complexes
pachytene - each pair of homologs (bivalent) coils tightly, crossing over occurs
diplotene - homologs separate but remain attached at chiasmata
diakinesis - separation of homolog pairs, chromosomes maximally condensed.
For females and meiosis II doesn’t finish till….
fertilization
What is nondisjunction?
failure of homologous chromosomes (MI) or sister chromatids to separate (MII)
What are the only three viable trisomies?
13, 18, 21
What is the only viable monosomy?
X (ie. Turner syndrome)
What is a Robertsonian translocation?
A translocation between two acrocentric chromatin which results in loss of the short arms of both chromosomes but does not affect the DNA content of the long arms. Most common is Der(13;14)
Paracentric vs pericentric inversions.
Paracentric inversions – breaks are in the same chromosome arm. Inverted segment does not include the centromere. acentric and dicentric chromosomes. If these gametes are fertilized they will not result in a viable conception. Carrier is at risk for multiple spontaneous miscarriages.
Pericentric inversions – breaks are in different arms. The inverted segment Includes the centromere.bnormal gametes with gain and loss of chromosome material. Carrier is at risk for multiple spontaneous miscarriages AND abnormal liveborn offspring.
How does FISH work?
A probe of interest (DNA strand of a known gene or locus of interest) is labelled with a fluorescent molecule. The probe is heat denatured to become single stranded. The patient metaphase spreads are also heat denatured. The probe and the target (patient’s chromosomes) are incubated together and the probe will anneal to the complimentary sequences on the chromosomes. The probe is visualized with a fluorescent microscope.
Can be done in interphase and metaphase.
How does microarray work?
A series of probes that map across the entire genome are arrayed on a microscope slide and hybridized simultaneously with equal amounts of patient DNA and control DNA. The patient and the control DNA compete for hybridization to the probes. If the DNA sequence is present in equal amounts in the patient and control there will be equal hybridization of patient and control. If a sequence is overrepresented in the patient (ex chromosome 21 sequences in a patient with Down Syndrome) then the patient DNA will out compete the control for hybridization. The ratio of patient and control hybridization is calculated to detect gains and losses in the genome.
Normal ratio = 0.8-1.2
Loss = <0.8
Gain = >1.2
Microarray analysis cannot detect balanced rearrangements, only unbalanced (gain or loss of DNA). Microarray also does not give information about mechanism and therefore recurrence risk. If gain of chromosome 21 DNA is detected it is not known if this is due to free trisomy 21 (+21) or due to the inheritance of an unbalanced Robertsonian translocation.
