Diagnostic Technologies - RM

Question 1

What technique hybridizes molecular probes to chromosomes?

Answer 1

FISH

Question 2

What are fragments of single stranded DNA homologous to region of DNA sequence of interest labeled with fluorochrome?

Answer 2

molecular probes

Question 3

What phase in the cell cycle is FISH performed at?

Answer 3

metaphase

Question 4

Why do you need an internal control in FISH? What does the control probe do?

Answer 4

to confirm hybridization is occuring (otherwise can’t tell if missing signal is due to disease related issue or technical error of probe not binding)
-control probe binds to different region of same chromosome that should be unaffected

Question 5

How many FISH signals indicates a normal individual?

Answer 5

2 signals per chromosome, 1 for the molecular test probe, 1 for the control probe indicating hybridization worked

Question 6

What are centromere probes used for?

Answer 6

chromosome enumeration (to see if there’s gain/loss)

Question 7

What are telomere probes used for?

Answer 7

confirms presence or absence of telomeric regions

Question 8

What does a single copy probe do?

Answer 8

identifies presence/absence of a gene or chromosomal rearrangment of interest specific to gene locus

Question 9

Why are subtelomere FISH probes important?

Answer 9

identifies very small (cryptic) deletions and rearrangments in DNA sequences in distal ends of chromsomes proximal to the telomeres that can’t be seen by standard karyotype
-3-5% of unexplained mental retardation due to cryptic terminal deletions

Question 10

What does chromosome painting do?

Answer 10

probes for many DNA fragments along chromosome make entire chromosome fluoresce so you can identify complex rearrangements and translocations due to different colors in a chromosome

Question 11

What is multicolor FISH good for detecting?

Answer 11

chromosomal translocations, duplications, deletions in multiple chromosomes using 1 hybridization

Question 12

What can’t multicolor FISH identify?

Answer 12

inversions, small deletions or duplications in chromosome

Question 13

What is the “critical region” in FISH?

Answer 13

portion of genetic anomaly that is always or almost always altered in mutational process
-what the probe is based off of since it can’t cover the entire deletion

Question 14

What are the disadvantages of FISH?

Answer 14

can’t detect diseases that FISH probe isn’t designated for, can’t screen all chromosomes or all loci

Question 15

What may be associated with subtelomeric microdeletion?

Answer 15

developmental delay

Question 16

What are contigous gene syndromes? What technique is commonly used to detect it?

Answer 16

deletions in regions in genome with clusters of closely associated genes whose normal functions are generally unrelated, causing multiple phenotypic anomalies
FISH used to detect these microdeletions

Question 17

What does WAGR stand for? What chromosome and arm of the chromosome is it on?

Answer 17

Wilms tumor, aniridia, genitourinary defects, mental retardation
11p

Question 18

What syndrome results from deletion of elastin gene on proximal long arm of 7? What are the features of it?

Answer 18

Williams syndrome
-coarse hair/skin, lack of flexibility in aorta, supravalvular aortic stenosis, skeletal and joint limitations, renal anomalies, excellent music skills, bad at math, blue sclera, stellate iris

Question 19

What is caused by 3 MB deletion on 22q?

Answer 19

VCFS

Question 20

What symptoms are associated with VCFS?

Answer 20

learning disability, hypotonia, short stature, cleft lip/palate, facial anomalies, cardiac anomalies (conotruncal heart defects), feeding difficulties at birth, weak immune system

Question 21

How is presentation of VCFS so variable?

Answer 21

• depends on complement of alleles in homologous chromsome (may be able to compensate for deletion)
• new combo of alleles for normal chromosome may cause different phenotype
• deletions can range in size and genes deleted

Question 22

What compares relative amounts of DNA from test source and reference source with known genetic complement?

Answer 22

microarray

Question 23

What does it mean if there’s an excess of test DNA signal in microarray compared to reference DNA signal?

Answer 23

duplication in the test DNA

Question 24

What does it mean if there’s an excess of reference DNA signal in microarray compared to test DNA signal?

Answer 24

deletion in test DNA

Question 25

What do gene arrays identify? What can’t they identify?

Answer 25

genetic polymorphisms, specific mutations, copy number variation (duplication/deletion)
-can’t identify balanced rearrangements

Question 26

What do expression arrays identify? How is it visualized?

Answer 26

• what DNA is actually being expressed in particular cells

- visualized as heat map of high level of expression/low level expression/median level expression based on colors

Question 27

Why are expression arrays important for cancer studies?

Answer 27

• expression of genes in tumors were assayed and upregulated genes for each tumor type were logged as the “fingerprint” for that type
• established clinical test to diagnose tumor type by running this panel of standards against the test specimen

Question 28

What do peaks in chromosome microarray results reveal? valleys?

Answer 28

peaks–>gain of segments–>duplications

valleys–>loss of segments–>deletions

Question 29

What is the best technique for cases of unexplained developmental delay, intellectual disability, autism spectrum disorders, and multiple congenital anomalies?

Answer 29

microarray

Question 30

What is the best for genome wide, relatively large numerical and structural abnormality testing?

Answer 30

karyotype analysis

Question 31

What is molecular diagnostics best for?

Answer 31

well defined, specific, very small (1-300 base pair) mutations
-targeted testing

Question 32

What size mutation is FISH best for detecting?

Answer 32

10 kb - 10 MB

Question 33

What can’t microarrays detect?

Answer 33

balanced rearrangements

Question 34

What two techniques are best for detecting mosaicism?

Answer 34

FISH, microarray

Question 35

What is best at detecting cosanguinety/identity by descent?

Answer 35

microarray

Question 36

If clinical findings suggest KNOWN numerical or structural disorder, what techniques are best?

Answer 36

FISH or karyotype (specific and targeted testing)