Brainscape's Knowledge GenomeTM

Browse over 1 million classes created by top students, professors, publishers, and experts.


See full index

Parasitology Lecture > ELECTROPHORESIS > Flashcards

ELECTROPHORESIS Flashcards

1
Q

is the movement of molecules by (DNA or RNA) by (applying specific voltage) an electric current

A

Electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

This can occur in air or solution or in a matrix to limit migration and contain the migrating material

A

Electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

True or False

Electrophoresis is commonly applied to the analysis of nucleic acids and proteins molecules

A

T

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

True or False

Nucleic Acids are mostly positive charged charged

A

F

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

Each phosphate group on a nucleic acid polymer is ________ making the molecule negatively charged

A

ionized

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

Under an electric current, DNA/RNA will migrate toward the?

A

positive pole (anode)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

In a matrix of agarose or polyacrylamide, migration under the pull of the current is impeded, it depends on the? (2)

A

size of the molecules

the spaces in the gel matrix

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

Things to note of Nucleic Acids in Electrophoresis (3)

A

It is affected by the size and the charge of the
particle

The nucleic acid will move toward the positive pole
(anode) because nucleic acids are negatively
charged.

From cathode it will migrate to anode

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

When phosphate group is of course ionized, it will be (1)___________ charged, so that would confer a (2) __________to your DNA and RNA.

A
  1. Positively
  2. Negativity
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

provide resistance to the movement of molecules under the force of an electric current

A

THE GEL SYSTEM

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

Gel system prevent diffusion and reduce convection
currents so that the separated molecules form
a defined group, or called as?

A

band

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

can then serve as a support medium for analysis of
the separated components

A

gel

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

The best gel matrix would be? (3)

A

unaffected by electrophoresis

simple to prepare

amenable to modification.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

The 2 best gel matrix in the analysis of NA in Electrophoresis

A

Agarose and Polyacrylamide

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

Agarose Gel

The size of the DNA is affected by its?

A

concentration

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

Agarose Gel
High concentration : ?

Low concentration : ?

A

impede migration

weak gel

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

Small pieces of DNA (50 to 500 base pairs [bp]) are resolved
on?

A

Higher agarose concentrations (ex. 2% to 3%)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

Larger fragments of DNA (2,000 to 50,000) are best resolved
in?

A

lower agarose concentrations (ex. 0.5% to 1%)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

The gel strength of any concentration of agarose will also? (2)

A

decrease over time

with exposure to chaotropic agents such as urea.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

DRAWBACK of Agarose gel Concentrations?

A

Agarose concentrations above 5% and below 0.5% are not practical.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

Identify what type of Electrophoresis.
-consists of very large pieces (50,00 to 250,000 + bp) of DNA
-Used Bacterial typing for epidemiological purposes

A

Pulse-field Gel Electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

Pulse-field Gel Electrophoresis.
For these very large DNA molecules, pulses of current applied to the gel in what dimension?

A

alternating dimensions to enhance migration

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

The four approaches of Pulse-field Gel Electrophoresis.

A

FIGE
TAFE
RGE
CHEF

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

The simplest approach to PFGE is?

A

field-inversion gel electrophoresis (FIGE) .

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
Q

What approach is this in PFGE?
-works by alternating the positive and negative
electrodes during electrophoresis
-
In this type of separation, the DNA goes
periodically forward and backward

A

field-inversion gel electrophoresis (FIGE) .

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
26
Q

is used for applications that require the resolution of chromosome-sized fragments of DNA, such as in bacterial typing for epidemiological purposes.

A

Alternating-field electrophoresis

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
27
Q

will yield a set of fragments that produce a band pattern specific to each type of organism. By comparing band patterns, the similarity of organisms isolated from various sources can be assessed. This information is especially useful in determining the epidemiology of infectious diseases.

A

Enzymatic digestion of genomic DNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
28
Q
  • Require a catalysts
  • Higher resolution capability for smaller
    fragments
A

Polyacrylamide Gels

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
29
Q

catalysts of Polyacrylamide Gels (3)

A
  1. ammonium persulfate (APS)
  2. N,N,N′,N′-tetramethylethylenediamine (TEMED)
    or 3. light activation
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
30
Q

Acrylamide, in combination with the cross-linker ___________________, polymerizes into a matrix that has consistent resolution characteristics

A

methylene bisacrylamide

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
31
Q

____________ is a natural polymer from living organisms?

_____________ is a synthetic material

A

agarose

polyacrylamide

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
32
Q

This gel allows precise control of the polymer properties and higher resolution.

A

polyacrylamide

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
33
Q

how does agarose gel polymerize?

A

cooling

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
34
Q

how does polyacrylamide gel polymerize?

A

polymerizations requiring a catalyst (APS + TEMED/Light Activation)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
35
Q

_________ produces free oxygen radicals in the presence of TEMED to drive the polymerization mechanism

A

APS

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
36
Q

What catalyst is this?

Free radicals are generated by a photochemical process using riboflavin plus TEMED.

A

Light activation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
37
Q

If Light activation is used:
Free radicals are generated by a photochemical process using?

A

riboflavin + TEMED

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
38
Q

If light activation is used, Excess oxygen inhibits the polymerization process.

Therefore ________or ____________ of the gel solution is done before the addition of the nucleating agents.

A

de-aeration or the removal of air

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
39
Q

The main advantage of polyacrylamide over
agarose is??

A

higher resolution capability of polyacrylamide for small fragments.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
40
Q

With single-base resolution, polyacrylamide
gels are used for: (4)

A

-nucleic acid sequencing
-mutation analyses
-nuclease protection assays
-other applications requiring high resolution of nucleic acids

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
41
Q

What type of electrophoresis?

-separates particles by size and charge
-Size and charge (charge/mass ratio)
-Increased sensitivity and immediate detection

A

CAPILLARY ELECTROPHORESIS

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
42
Q

CAPILLARY ELECTROPHORESIS
size (small, ____migration; large, ____migration)

charge (________, fast migration; ________, slow migration)

A

size (small, fast migration; large, slow migration)

charge (negative, fast migration; positive, slow migration)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
43
Q

CAPILLARY ELECTROPHORESIS
Because the size and charge of DNA work counter to each other, a __________ in the capillary will resolve the DNA fragments mostly according to size.

A

polymer (gel)

-so bale more on SIZE ang capillary electrophoresis.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
44
Q

CAPILLARY ELECTROPHORESIS
Negatively charged molecules are completely (1) _________ at ______pH, whereas positively charged solutes are completely
(2)_________ in ____-pH buffers

A

ionized, high

protonated, low

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
45
Q

True or False

In Capillary Electrophoresis, Nucleic acids do not separate well in solution.

A

T

(As the size or length of a nucleic acid increases (slowing migration), so does its negative charge (speeding migration),
effectively confounding the charge/mass resolution

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
46
Q

In Capillary Electrophoresis, It is VERY important that the nucleic acids are??

A

completely denatured (single stranded) so that it will be
separated according to its size because the secondary
structure (dsDNA) will affect the migration speed.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
47
Q

Compared with traditional slab gel electrophoresis, the
capillary system has the advantages of?

A

Increased sensitivity and immediate detection

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
48
Q

to carry the current and protect the samples during electrophoresis

A

BUFFER SYSTEM

49
Q

Tris-buffers
Most common NA?

A

DNA

50
Q

Types of Tris-Buffers (3)

A
  1. Tris-borate EDTA (TBE)
  2. Tris phosphate EDTA (TPE)
  3. Tris acetate EDTA (TAE)
51
Q

Basaha ra (Tris-Buffers)

A

TBE has a greater buffering capacity than TAE. Although the
ion species in TAE are more easily exhausted during
extended or high-voltage electrophoresis, DNA will migrate
twice as fast in TAE than in TBE in a constant current.

52
Q

modify sample molecules in ways that affect their migration

A

Buffer Additives

53
Q

Examples of these buffer additives are: (3)

A
  1. Formamide
  2. Urea
  3. Detergents
54
Q

Denaturing agents that break hydrogen bonds between complementary strands or within the same strand of DNA or RNA (2)

A

(Formamide & Urea)

55
Q

___________ and heat added to DNA and RNA break and
block the hydrogen-bonding sites, hindering complementary sequences from reannealing. As a result, the molecules become long, straight, unpaired chains.

A

Formamide

note. block the hydrogen-bonding sites, no anneal, and long, straight, unpaired chains

56
Q

________ and heat in the gel systems maintain this conformation such that intrachain hybridization (folding) of the nucleic acid molecules does not affect migration speeds, and separation occurs strictly according to the size or length of the molecule.

A

Urea

note. intrachain hybridization (folding) and strict separation to size and length

57
Q

RNA Denaturing Agents that reacts with amino groups on the RNA to prevent base pairing between complementary nucleotides and with aldehydes (e.g., formaldehyde, glyoxal) which also disrupt base pairing.

A

methylmercuric hydroxide (MMH)

58
Q

ELECTROPHORESIS EQUIPMENT
-are run in acrylic gel boxes or baths that are divided into two parts, with a platform in the middle on which the gel rests

A

Horizontal gels

59
Q

ELECTROPHORESIS EQUIPMENT
What colored connector is attached to positive port? (1)
What colored connector is attached to negative port? (2)

A

RED
BLACK

60
Q

ELECTROPHORESIS EQUIPMENT
NA will migrate toward the?

A

Positive (red) pole

61
Q

ELECTROPHORESIS EQUIPMENT
-placed across the box at each end of the bath compartments are connected to a power supply through the walls of the container

A

The electrodes (platinum wires)

62
Q

ELECTROPHORESIS EQUIPMENT
-is submerged in electrophoresis buffer that fills both compartments and makes a continuous system through which the current flows.

A

Gel

63
Q

ELECTROPHORESIS EQUIPMENT

The THICKNESS OF THE GEL and THE VOLUME OF THE BUFFER affect the _________, and therefore the migration of the sample, so these parameters are kept ________ for consistent results.

A

current

constant

64
Q

ELECTROPHORESIS EQUIPMENT
Because the gel is submerged throughout the loading and electrophoresis process, horizontal gels are sometimes referred to as ??

A

submarine gels

65
Q

The volume of the gel solution will determine the?

A

thickness of the gel.

66
Q

True or False
Gel is mixed with agarose and Polyacrylamide

A

F

Gel is mixed with buffer

67
Q

Molten agarose is cooled to between what temp?

A

55C and 65C

68
Q

inserted into the top of the gel to create holes, or wells, in the gel into which the sample will be loaded

A

Comb

69
Q

Comb
Size of the teeth = ?
# of the teeth = ?

A

capacity

of wells

70
Q

Two types of combs used in polyacrylamide electrophoresis

A
  1. Regular combs- horizontal gel
  2. Sharkstooth combs - vertical gel
71
Q

Combs
________ gels are cast between glass plates that are separated by spacers.

A

vertical gel (Sharkstooth combs)

72
Q

Combs
The spacers determine the thickness of the gel, ranging from _____ to ______mm.

A

0.05 to 4 mm.

73
Q

Combs
The bottom of the gel is secured by _____or by a ______ in specially designed gel casting trays.

A

tape or by a gasket

74
Q

Combs
After the addition of catalyst and nucleating agents, the (1) ___________ is poured or forced between the glass plates with a (2) _______ or a (3) _______

A

1 Liquid acrylamide
2 Pipet
3 Syringe

75
Q

Combs
For light-activated polymerization, the gel between the glass plates is exposed to a _________

A

light source.

76
Q

Comb
It is important to prevent air from getting into the gel or beneath the comb. ________ will form discontinuities in the gel, and oxygen will inhibit the polymerization of the acrylamide.

A

Bubbles

77
Q

The comb is of a thickness equal to that of the
________ so that the gel will be the same
thickness throughout.

A

spacers

78
Q

True or False
Combs are only used in Polyacrylamide Gel Electrophoresis and not on Agarose

A

F

79
Q

Combs
for vertical gels/Sharkstooth combs, its usually ____________

horizontal/Regular combs, could either be _________ or _________

A

polyacrylamide

agarose or polyacrylamide gels

80
Q

Gel Loading consists of (2)

A

Tracking dye
Density agent

81
Q

used to monitor the progress of the electrophoresis run

A

Tracking dye

82
Q

The dyes migrate at specific speeds in a given gel concentration and usually run ahead of the SMALLEST fragments of DNA

A

Tracking dye

83
Q

3 ex. of Density Agents

A

Ficoll
sucrose
glycerol

84
Q

increases the density of the sample as compared with the electrophoresis buffer.

A

Density Agents

85
Q

DAWBI?

A

kamo nay memo sa number2 sa tracking dye uy kapoy

86
Q

Two Most Common Tracking Dye

A

Bromophenol blue and Xylene cyanol green

87
Q

is a tracking dye that is used for many applications

A

Bromophenol blue

88
Q

is another of the chromophores used as tracking dyes for both agarose and polyacrylamide gels

A

Xylene cyanol green

89
Q

True or False
Tracking dyes are associated with the sample DNA, and thus they affect the separation.

A

F
Tracking dyes are not associated with the sample DNA, and thus they do not affect the separation.

90
Q

DETECTING SYSTEM (2)

A

Fluorescent Dyes
Silver Stain

91
Q

The agents used most frequently for visualization of bands
after electrophoresis are (2)

A

Fluorescent Dyes
Silver Stain

92
Q

an intercalating agent and most widely used dye in EARLY dna and rna analyses.

A

Ethidium bromide

93
Q

EtBr Under excitattion with UV light (nm)?

A

300 nm

94
Q

EtBr in DNA emits visible light at (nm)?

A

590 nm

95
Q

DNA separated in agarose or acrylamide and exposed to EtBr will emit ______ light when illuminated at “300 nm”

A

orange

96
Q

After electrophoresis, the agarose or acrylamide gel is soaked in a solution of ____ to _____-mg/mL EtBr in running buffer (TAE, TBE, or TPE) or in TE.

A

0.1- to 1-mg/mL EtBr

97
Q

ETBR
After soaking or running in dye, the DNA illuminated with UV light (300nm) will appear as ________ bands in the gel.

A

orange

98
Q

Minor Groove-Binding Dyes (2)

A

SYBR green
SYBR gold

99
Q

SYBR green I - ?
SYBR green II - ?

A

SYBR green I - dsDNA
SYBR green II - ssDNA, RNA

100
Q

SYBR green I differs from EtBr in that it?

A

it does not intercalate between bases; it sits in the minor groove of the double helix.

101
Q

SYBR green in association with DNA or RNA also emits (visible) light in the orange range (nm)?

A

522 nm

102
Q

In agarose gel electrophoresis, SYBR staining is __ to ___ times more sensitive than EtBr

A

25 to 100

so mas SENSITIVE si SYBR kaysas Etibromide nato

103
Q

Fluorescent Dyes
_________ can also be added directly to the DNA sample
before electrophoresis

A

SYBR green

so SYBR ma add na b4 electrophoresis unlike sato ETBR nga after pa kay mag soak2 paman nasya

104
Q

but in SYBR, basaha lang

A

DNA prestaining decreases the amount of dye required for
DNA visualization but lowers the sensitivity of detection and
may, at higher DNA concentrations, interfere with DNA
migration through the gel.

105
Q

True or False
Because SYBR green is not an intercalating agent, it is mutagenic

A

F
Because SYBR green is not an intercalating agent, it is not as mutagenic

106
Q

True or False
SYBR green is safer to use than EtBr

A

T

107
Q

True or False
Ethidium bromide is good for Real time PCR

A

F
SYBR green

108
Q

What is this type of fluorescence dye?
-fluorescence increases more than 1,000- fold upon binding to double- or single-stranded DNA or to RNA

A

SYBR Gold

109
Q

Like SYBR green, SYBR gold is excited by UV
light (nm)?

A

300 nm

110
Q

SYBR gold emits (visible) light at (nm)?

A

537 nm

111
Q

Another sensitive staining system originally developed for protein visualization, increased sensitivity.

A

Silver Stain

112
Q

Silver Stain consists of 2 procedures?

A

Silver diamine (ammoniacal silver)
Silver nitrate

113
Q

Silver Stain
After electrophoresis, the sample is fixed with (2)

A

methanol
acetic acid

114
Q

Silver Stain
The gel is then impregnated with ammoniacal
silver (silver diamine) solutions or silver nitrate in a ___________ solution.

A

weakly acid

115
Q

Silver Stain
Interaction of silver ions with acidic or nucleophilic groups on the target results in _________ or ______________ of metallic silver under optimal pH conditions.

A

crystallization or deposition of metallic silver

116
Q

Silver Stain
The insoluble black silver salt precipitates upon
introduction of ________ in a weak acid solution, or alkaline solution for ____________

A

formaldehyde (weak acid sol’n)
silver nitrate (alkaline sol’n)

117
Q

Silver Stain
____________ is best for thick gels

____________ is considered to be more stable

A

silver diamine

silver nitrate

118
Q

These are especially useful for protein analysis
and for detection of limiting amounts of product.

A

Silver Staining

Parasitology Lecture (1 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2024 Bold Learning Solutions. Terms and Conditions