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Flashcards in Molecular Diagnostics - 3 lectures Deck (76):
1

difference between direct and indirect methods

indirect - looking at markers that tend to be close to disease allele (think recombination)

direct - everything else

2

purpose of linkage analysis

to find a genetic loci that is associated with a disease

3

what phase does crossover happen in the DNA

prophase of meiosis I

4

which loci experience crossover more - ones that are farther apart or closer together

farther apart

5

marker that is usually inherited with the disease causing mutation

polymorphic marker

6

genes or marker that are not close together or on different chromosome experience what percentage of recombination requency

50% recombination frequency (independent assortment) and are considered not linked

7

genes that are close together experience what percentage of recombination frequency

less than 50% and are considered linked

8

particular arrangement of a chromosome that is inherited together

haplotypes

9

low or high frequency of recombination - two loci that are close together

LOW

10

how are disease causing gene actually identified

sequencing nearby regions of the genome or using human genome data

11

restriction enzymes cutting at palindromic sequences is used in what method

PCR RFLP analysis

12

a disease individual without the marker being used or a non affected person with the diseased marker is a result of?

recombination

13

challenges to linkage analysis

recombination and loci heterogenity(same phenotype but different loci)

14

methods that query the whole gene

karyotyping (G banding), array CGH, SNP chip, expression arrays, special karyotyping, and next gen sequence

15

significance of a single bp difference every 1000bp

digestions of DNA from different individuals will result in different patterns of DNA fragment

16

what happens at restriction sites that have single bp change

they are destroyed

17

RFLP data was originally obtained from what other method

southern blotting

18

disadvantage of southern blotting

lengthy procedure, not automated, radioactivity, expensive

19

as of now RFLP data is obtained using what?

PCR which uses restriction enzymes

20

Direct RFLP used to diagnose what illness

sickle cell anemia

21

data for direct RFLP depends on what?

locus and restriction enzyme

22

ASO used for diagnoses of what diseases

cystic fibrosis and hemochromatosis

23

when do ASO become less useful

as the disorder exhibits more allelic or genetic heterogenity

24

ASO bind to what

single allele of a gene - only possible if exact mutation has been identified

25

what are the triple repeat disorders and what method is used to detect them

fragile X, huntingtons, myotonic dystrophy

PCR

26

how can PCR tell you it is triple repeat disorder

the size of the PCR product - the genotype will have a lot more than normal repeats

27

how will the PCR product of a person with triple repeat disorder show on the agarose gel

the DNA wouldn't travel as far as the normal one because it is too big

28

PCR can be used to detect

Duchenne because it is a deletion

29

limitations to PCR

difficult to optimize so can't detect carriers

30

can be used to detect aneuploidy, insertions, deletion, large translocations

karyotyping (G banding)

31

disadvantages to karyotyping

only detects large alterations (deletion >5kb or even more) and has low resolution

32

in G banding, the dark and light bands represent what

dark: A-T rich region
light: C-G rich region

33

keypoint of FISH

resolution is improved over G banding

34

how long are the probes in FISH

10-100kB

35

difference between the two types of FISH (SKY FISH and chromosome specific)

SKY FISH - paints the whole chromosome a specific color

chromosome specific - hybridizes to a specific locus on the chromosome (only one locus)

36

DiGeorge's Syndrome or 22q11.2 is detected using what method

FISH

can also use G banding but might not get great results

37

what is abnormal in digeorges

pharyngeal arch development

38

symptoms of digeorge's

CATCH22
cardiac anomaly most important

39

advantage of FISH

better resolution of G banding

40

disadvantage of FISH

have to know exactly what you are looking for

41

FISH can be used to detect what disorder

trisomy

42

SKY FISH most importantly used to detect

translocations since it colors each chromosome a specific color

43

disadvantage of SKY FISH

can't detect small deletions since whole chromosome is painted one color

44

if you suspect a patient of having digeorge's and you do FISH method and you have a red signal in both chromosome 22s, what doest that mean

patient does not have digeorge's since red signal is present in both chromosomes - would have been digeorge's if red signal only present in one chromosome

45

advantage of array CGH

can detect copy number changes in DNA throughout the genome, genome amplification, deletions (CNV - copy number variant)

46

disadvantage of array CGH

does not detect balanced rearrangements

47

array CGH detect deletions and insertion at very high or very low resolution?

high resolution

48

what method can detect gain or loss of DNA copies

array CGH

49

why cant array CGH detect Philadelphia chromosome (translocation between 9 and 22)

it can only detect additions and deletions of chromosomes and not balanced translocations

50

why can SKY FISH detect Philadelphia chromosome (translocation between 9 and 22)

because each chromosome is painted a different color so it will show chromosomes having two different colors in this case

51

FISH can detect deletions at about how bp levels

~100,000 bp

52

involves labeling genomic DNA fragments and allowing them to hybridize to DNA spots adhered onto a microarray slide where each spot represents various alleles of a gene.

SNP microarray chips

53

advantage of SNP

identify hundreds of thousands of polymorphic sites in a single experiment

54

CYP450 is used for?

phase I metabolism of about 20% of prescribed drugs

55

can a person's allele affects how drug is metabolized

yes

56

DNA is isolated from a patient who was born with congenital malformation of several different organs. The patient’s DNA was broken into small pieces, labeled, and then compared to an equal amount of a reference standard by hybridization to an array of DNA probes that represent sequential regions of the genome separated by 50 kb.

what technique is this?

array CGH - we are comparing to a reference

57

A patient is identified to have Prader Willi syndrome as a result of uniparental disomy. In this case, he has lost the paternal contribution of chromosome 15 and instead he has two copies of his maternal chromosome 15 which are sister chromatids (the non-disjunction occurred in meiosis 2). Which technology would best diagnose this condition?

SNP chip

58

microarray that contains features representing a set of known expressed (transcribed) sequences

cDNA microarray

59

what can cDNA be used for

May be used to identify a set of genes or all genes that are expressed in a cell or tissue

May be used to compare gene expression between cell or tissue samples

60

are prices of sequencing genome going up or down

down

61

a method to obtain the base pair sequence of a piece of DNA

Sanger (dideoxy) sequencing

62

what does the dideoxynucleotide do to the DNA strand in sanger sequencing

it terminates the synthesis of the DNA strand by making sure the free 3' OH is absent

63

current method of sequencing uses what

fluorescence label and capillary electrophoresis

64

how can one tell the position of the base relative to the primer and the type of base in sequencing

position of the base - length of DN sequence
type of base - color of the DNA

65

A patient is suspected to have a genetic cause associated with his symptoms. A test is given which detects a deletion of 500,000 basepairs of DNA from his chromosome 7. What test was used to detect this?

array CGH



could also use FISH - since you know exactly what you are asking for

66

what happens if you sequence a gene and you find a base pair that is altered on the second run, what do you infer?

could be a mistake or not so you run the sequence many more times

67

what happens if you sequence a gene and for a particular bp, half of the time, the bp shows T and the other half it shows as G. what do you infer?

polymorphism or mutation

68

analysis that can be used to detect and quantify the amount of a particular protein

western blotting

69

detection using a labeled antibody system (typically an enzyme label and a chemiluminescent substrate)

western blotting

70

methods that only focus on one gene

southern, northern, western, sequencing of PCR product, ASO detection on DNA, allele specific primers for PCR

71

Techniques that can detect only what is asked for

ASO blot, PCR, FISH, southern

72

point mutation that does not change the AA code

silent mutation

73

point mutation that changes the protein

missense mutation

74

point mutation that creates a stop code

nonsense mutation

75

mutation that changes base from one purine to another purine, or one pyrimidine to another pyrimidine

transition mutation

76

mutation when a purine is replaced by a pyrimidine or vise versa

transversion mutation (less common)