Lecture 4: Diagnostic Tech's Flashcards
How does FISH work? What’s the most common goal of it?
Well-char’d and locus & chromosome specific Molecular probes (ie a fragment of DNA) hybridizes to chromosomes. To detm if a gene, a specific mutation, or a particular chrom rearr is present or absent. A probe only detects the site from which it was derived.
Repeat sequence probes are usually isolated from what regions?
Telomere or centromere regions.
A unique sequence, or single copy, probe is usually isolated from where, and for what purpose?
Cloned DNA of a disease-causing gene or fragment of DNA of known location assocd with a particular gene. Used to ID the presence or absence of the gene, gene region, or chrom rearr of interest.
A deletion at the ends of chrom’s are assocd with many diff disorders due to the abundance of coding genes in this region. What type of FISH allows us to ID cryptic deletions and rearr’s that cannot be seen by std karyotype analysis, which account for 3-5% of unexplained MR?
Subtelomere FISH.
Chromosome painting, or Whole Chrom Paint, is a cocktail of many unique DNA frags from along the whole length of a C such that following hyb, the whole thing fluoresces. This type of probe is most useful in IDing what?
Cpx rearr’s or marker chrom’s, say a chrom with extra material from God knows where. This can help ID the source. It’s most useful when looking at translo’s. For ex, could visulaize a 3:1 segregation, resulting from the parent probably being a balanced t carrier.
What is type of chromosome painting used to detect multiple C’s with one hyb? What’s it very good at detecting and not so good at?
Multicolor FISH. To see over 3 colors, a computer-assisted imaging system is required. Good at C rearr’s (t, large dels or dups), can’t ID inv’s, small dels or dups.
What’s the critical region that probes for FISH cover?
That portion of the genetic anomaly that is always or almost always altered during a given mutational process.
FISH is commonly used to detect microdeletions such as those seen in contiguous gene syndromes. What are these syndromes? List as many of these diseases as you can.
Deletion of region where clusters of genes exist that confers a particular clinical outcomes, even though indiv the functions of those genes are unrelated. WAGR-11p, Miller-Dieker/Lissencephaly-17p, Williams syn-7q, VCFS-22q, 1p-syndrome, and Prader Willi/Angelman syndromes. The phenotype depends on which genes are lost.
WAGR
Wilms tumor and aniridia, larger deletions add on GU defects and MR. 11p.
Williams syndrome
Deletion of the elastin gene on 7q, once considered just a microdeletion of a single gene (wrong). Devel problems, group homes, supravalvular aortic stenosis, renal anomalies, coarse skin/hair, jt and skel limits, awesome at music, stink at math, outgoing and friendly, blue sclera, stellate iris.
VCFS
Interstitial microdeletion of 3MB on C 22 involving 40 genes. Learning disabilities, short stature, cleft lip/palate, facial dysmorphologies, cardiac anomalies, feeding difficulty at birth, weak immune system.
Elaborate on the mechanism by which the 3MB deletion in VCFS comes about? Hint: repeat sequences and crossing over.
4 repeat sequences exist within the 3MB region, and bc of their similar sequences, it’s possible the wrong repeats will pair and if recomb occurs, unequal crossing over occurs resulting in the 3MB deletion with the reciprocal product a duplication.
What technique uses a machine to look at copy # variation after hybridizing test DNA to known reference DNA strand in order to assess what color dominates and hence a dup or del? Besides CNV what else can it identify? What will this assay not usually be able to detect?
Microarray-comparative genomic hyb. Genetic polymorphisms or specific mutations. Balanced chrom rearr’s cuz the total amt of DNA is conserved. Microarrays are a generalized genome screen for small to large mutations.
What type of array involved extracting RNA from a tissue of interest, making cDNA from it and labeling with a fluorochrome, which is then hybridized to the slide? By looking at the same gene in diff tissues, it’s possible to detm which genes are most imp in that tissue.
Expression arrays.
Instead of reading array data out per gene or allele, what type uses DNA fragments directly assoc’s with locations on chrom’s? It’s used regularly today.
Chromosome Microarray.
