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BIOL 2030 > Section 3 - Module 11 > Flashcards

Section 3 - Module 11 Flashcards

1
Q

What test is used for mutagenicity?

A

Ames test

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2
Q

What is Ames test?

A

Inclusions of rat liver enzymes to mimic the chemical modifications of potential mutagens in the human body

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3
Q

Discovery of Ames test?

A

Liver enzymes could make the chemical more or less mutagenic

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4
Q

Why do bacteriophages grow on some bacterial strains, but not others?

A

Because of ‘Type I” restriction endonucleases

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5
Q

What do “Type I” endonucleases do?

A

Recognize specific DNA sequences and cleave DNA

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6
Q

What type of restriction endonucleases are “restriction enzymes”?

A

Type II

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7
Q

What do “Type II” endonucleases do?

A

cleaves DNA within the recognition site

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8
Q

What type of restriction endonuclease is more useful in molecular biology?

A

Type II

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9
Q

The DNA restriction site is a __

A

palindrome

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10
Q

What is a palindromic sequences?

A

a sequence of nucleotide bases reads the same on the top strand as the sequence of nucleotide bases reads on the bottom strand of the DNA molecules in 5’ - 3’ direction

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11
Q

What is the resulting form of EcoRI restriction endonuclease?

A

“sticky” ends

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12
Q

1st endonuclease isolated?

A

EcoRI and BamHI

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13
Q

3rd endonuclease isolated?

A

HINDIII

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14
Q

What endonuclease is from the bacterium Escherichia coli?

A

EcoRI

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15
Q

What endonuclease is from the bacterium Bacillus amyloliquefaciens?

A

BamHI

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16
Q

What endonuclease is from the bacterium Haemophilus influenza?

A

HindIII

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17
Q

Why don’t bacterial restriction endonucleases attack the host’s own DNA?

A

Because the host methylates a base in every cop of the RE site within its own genome.

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18
Q

What can rejoin the DNA sequences cut by the Type II restriction endonuclease?

A

ligases, rejoining sticky ends

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19
Q

What is gel electrophoresis?

A

a method of sorting DNA (and RNA) sequence fragments by size. Uses electrical field to more charged DNA towards the positive electrode.

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20
Q

T/F. electrophoresis can occur in a liquid?

A

False. Electrophoresis must occur in a gel such as agarose

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21
Q

In gel electrophoresis what moves furthest towards the cathode?

A

Shorter molecules

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22
Q

What visualizes (stains) size-fractionated DNA?

A

DNA-binding florescent dye

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23
Q

What type of dye is EtBr?

A

intercalating (between bases)

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24
Q

Migration rate of a _____ DNA molecule is inversely related to log of its molecular mass (or # of base-pairs [bp]).

A

linear

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25
What can effect the migration of DNA in agarose gel matrix?
the concentration of agarose
26
As agarose concentration increases, pore size in gel matrix ___
decreases
27
What are the different topologies (conformations) of DNA molecules?
Linear, relaced circular, and supercoiled
28
T.F. The topology of DNA strands affect their rate of migration in gels?
True
29
In cells what is the charge of supercoiled DNA?
negative
30
What can speed up the migration rate of DNA fragments during agarose gel-electrophoresis?
greater voltage
31
What are some factors that DO NOT influence the rate of migration of DNA molecule's during agarose gel-electrophoresis?
The %GC content or sequence of a DNA molecule
32
Requirements for DNA synthesis in vitro?
1) a strand of DNA to act as a template 2) a short, single strand of DNA complementary to part of the template (the primer) 3) DNA polymerase 4) deoxyribonucleoside triphosphates (dNTPs) 5) Mg+ (needed by polymerase)
33
What direction does DNA synthesis always proceed?
3'
34
35
What does PCR stand for?
polymerase chain reaction
36
Mullis' insight (associated with PCR)
enzymatic copying of double-stranded DNA using 2 primers, complementary to opposite strands could lead to exponential increase in amount of target sequence
37
PCR required DNA to be cycled repeatedly through how many temperatures?
3
38
Generally how many cycles of PCR?
30-35 (in theory more than a billion-fold amplification)
39
Order of PCR steps
1) Denaturation 2) Annealing 3) Elongation or extension
40
Denaturation PCR step
94-96 C, dsDNA denatured to ssDNA
41
Annealing PCR step
50-65 C, primers bind to their complementary sequences, Tm is dependent on length and base composition of primers
42
Elongation or extension PCR step
72 C, DNA polymerase binds to the annealed primers and extends DNA at the 3' end of the chain
43
What are the 'ingredients" in PCR?
dinucleotide triphosphates, Mg2+, primers, template DNA, thermostable DNA polymerase, a salt, Tris (pH control), plus stabilizers
44
taq
Thermus aquaticus, a thermostable DNA polymerase
45
What are Dinucleoside triphosphates (dNTPs)
dATP, dCTP, dGTP, dTTP
46
What are primers in PCR?
short molecules of ssDNA (aka oligonucleotides),
47
What is the results of priming between two logos annealed to opposite strands?
gives exponential growth of product
48
What effects the size of the PCR products?
How far apart the annealing sites of the 2 primers
49
How long are primers typically?
18-25 bp (enough to have binding sites, but also not so long they cost a lot of energy)
50
Applications of PCR?
Amplifying target sequences for further study, detection of rare DNA sequences,
51
What is PCR not good at?
Determining abundance of the rare sequences
52
What type so growth does the early phases of PCR experientces?
exponetial
53
What type of growth does later cycles of PCR experience?
linear
54
What types of growth does end stage of PCR experience?
plateau
55
What phase provides the best information to estimate starting amount of DNA (or RNA) template in PCR?
Log-linear phase
56
Alternative name for real-time PCR
quantitative PCR (qPCR)
57
qPCR procedure
uses a reporter dye to monitor the PCR product
58
What dye is used most commonly in qPCR?
SYBR green
59
What grooves does SYBR fluorescence most commonly bind to ?
minor groove in dsDNA
60
Application of qPCR?
1) quantify amount of starting DNA of a particular sequence 2) measuring rate at which a particular gene is transcribed

BIOL 2030 (7 decks)

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