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MODX > NUCLEIC ACID AMPLIFICATION PART 2 > Flashcards

NUCLEIC ACID AMPLIFICATION PART 2 Flashcards

1
Q

difference of probe amplification to target amplification

A

number of targets nucleic acid sequences in the sample is not changed

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2
Q

purpose ofoligonucleotide

A

prime the dna polymerase

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3
Q

3 types of nucleic acid amplification technique

A

target, signal, probe

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4
Q

the amplified targets in dna sequence is called as

A

amplicons

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5
Q

short stretches of nucleic acids

A

probes

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6
Q

are synonymous to primer

A

probes

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7
Q

in probe amplification, what we are amplifiying is

A

probe itself

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8
Q

probes are attached to a fluorogenic dye such as

A

reported dye or guencher dye

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9
Q

__ detects particular infectious
organisms as well as gene rearrangements (ex. sickle
cell anemia)

A

Probe amplification

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10
Q

PROBE AMPLIFICATION

Can also detect chromosomal translocations (ex._____ ), and chromosomal breakages

A

down syndrome

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11
Q

Also used to detect changes in oncogenes (promotes
cancer development), and tumor suppressor genes

A

PROBE AMPLIFICATION

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12
Q

probe amplification is Also used to detect diagnosis of inherited disorders (ex.
_______

A

cystic fibrosis

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13
Q

Also used to detect carrier status, polymorphic markers,
and establish identity or non-identity

A

probe amplification

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14
Q

Also used to aid in donor selections and confirming an
organism isolated in the culture

A

probe amplification

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15
Q

probe amplification is used to confirm what organism

A

mycobacterium legionella
salmonella
E coli (diarrheagenic e.coli)
campylobacter

fungi
blastomyces dermatitidis
coccidiodes imitis
cryptococcus neoformans
histoplasma capsulatum

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16
Q

probe amplification

Can also detect agents of sexually transmitted disorder
such a

A

Chlamydia trachomatis,
Neisseria gonorrhoeae, cytomegalovirus, Epstein Barr Virus,
herpes simplex virus,
and detection of viral loads of HIV and HCV

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17
Q

3 major procedures of probe amplification

A

ligase chain reaction
strand displacement amplification
qb replicase

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18
Q

ligase chain reaction is manufactured and marketed by

A

abbott

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19
Q

strand displacement amplification is manufactured and marketed by

A

beckton dickinson

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20
Q

qb replicase is manufactured and marketed by

A

vysis

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21
Q

PCR and probe amplification is similar, however, there’s one step that is added in probe amplification.

A

ligation - connecting probe together using ligase enzymes and that probe will automatically analyzed and amplified

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22
Q

In LCR, __ is used to
ligate the adjacent primers
together

A

DNA ligase

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23
Q

a method for amplifying synthethic primers/probes complementary to target nucleic acid

A

ligase chain reaction

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24
Q

in LCR, the ligated primers can serve as ___ for annealing and ligation of additional primers

A

template

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25
Q

product of LCR is

A

ligated primer thus classifying it as a probe amplification

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26
Q

Lcr can be used to detect point-mutation in a target sequence

A

the point mutation that occurs in B globulin of patients with SICKLE CELL DISEASE is one of the first applications for LCR

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27
Q

SDA or strand displacement amplification is an example of what process

A

isothermal amplification process

28
Q

isothermal amplification process means there will no changed in what parameter

A

no changes on temperature

29
Q

STRAND DISPLACEMENT AMPLIFICATION is Originally developed and patented by

A

Beckton dickinson in new jersey year 1991

30
Q

primers of strand displacement amplification is incorporated to a specific __

A

restriction enzyme

31
Q

strand displacement amplification creates a __ using restriction endonuclease enzyme

A

nick

32
Q

target in strand displacement amplification si

A

double stranded dna

33
Q

DNApolymerase will elongate the

A

dATPase

34
Q

two stages to SDA process

A

◆ Target Generation Phase
◆ Exponential Probe or Target Amplification Phase

35
Q

Target DNA is denatured by heating to ______°C

A

95

36
Q

which stage of SDA is this

A

at each stage end of the target sequence, a primer and a probe bind close to each other

37
Q

the probe will have a site or a recognition sequence specific for ___

A

restriction enzyme

38
Q

the modified nucleotide (dATPas) will be elongated and the ____ will extend the primers incorporating to a modified nucleotide

A

Exonuclease-deficient DNA polymerase derived from E. coli

39
Q

a stage of SDA in which the product of this amplification is millions of copies of the initial probe

A

exponential probe/target amplification phase

40
Q

application of SDA

A

detection of M. Tuberculosis
C. trachomatis
N. gonorrhea

41
Q

In SDA, addition of a ___ to the reaction produces a fluorescent signal that corresponds to the amount of amplified target

A

fluorogenic probe

42
Q

method for amplifying probes that have specificity for a target sequence

A

QB replicase

43
Q

QB replicase is named for the major enzyme that is used to amplify probe sequences

true or false

A

true

44
Q

the RNA dependent RNA polymerase/ QB replicase came from

A

bacteriophage QB

45
Q

2 types of probes that is used by QB replicase

A

reporter probe
capture probes

46
Q

4 steps of QB replicase methodology

A

hybridization, capture, wash, and release

47
Q

how many hybridization procedure are done in QB replicase

A

2 hybridization

48
Q

steps in QB REPLICASE

A

capture probe A - will capture target RNA in hybridization connected to magnetic bead. Upon connecting to magnetic bead, it will under go capture, washing, and release. Once release, it will go under hybridization again using capture B. Will under go capture, washing, and release again.

It will become purified reported probe, this reporter probe will be act upon by the QB replicase enzyme in order to amplify or target this reporter probe.

These reporter probe will have the specificity for the target sequence because of the so called MIDVARIANT 1

49
Q

Reporter probes–RNA molecules that have specificity for the target sequence and also contain a promoter sequence that is recognized by the QB replicase

A

midivariant-1

50
Q

the qb replication is very efficient with the generation of _____ RNA molecules/probes in less than 15 mns

A

10^6-10^9 RNA

51
Q

application of QWB replicase

A

detection of infection organisms such as
mycobacteria
chlamydia
HIV
CMV

and genetic disorders such as
sickle cell anemia, cystic fibrosis, huntington’s disease

duchenne muscular dystrophy
vwillebrand diseases

52
Q

signal amplification will amplify

A

signals that are attached to targets

53
Q

signal are attached to labeled enzymes, and used upon by the technique called

A

chemilluminescence

54
Q

chemilluminescence is measured using

A

luminometer

55
Q

applications of signal amplification

A

designed to increase signal strength by increasing concentrations of the label

56
Q

labels example in signal amplification

A

enzyme label or antibody labels

57
Q

signal amplification is used in detection of what organisms

A

HCV, HIV, Legionella

58
Q

4 methods of signal amplification

A

branched DNA amplification
hydrid capture assay
cleavage-based amplification
cycling probe

59
Q

branched DNA amplification is developed by

A

chiron corp

60
Q

target nucleic acid of branched dna amplification is can be

A

DNA or RNA

61
Q

series of ____ are used to capture to target nucleic acid and then additional extended probes bind to target nucleic acid, and then to multiple reporter molecule, loading the tarhet nucleic acid with isgnal

A

short oligomer probes

62
Q

enzyme used in branched DNA amplification is

A

alkaline phosphatase

63
Q

the target nucleic acid is released from the cells, the DNA is denatured if DNA is the target, and the target nucleic acid binds to capture probes that are fixed to a solid support

A

branched dna amplification

64
Q

generations of branched DNA amplification

A

first, second, and third

65
Q

substrate used for the alkaline phosphatase

A

dioxetane

66
Q
A

MODX (17 decks)

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