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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
Q

What can effect the migration of DNA in agarose gel matrix?

A

the concentration of agarose

26
Q

As agarose concentration increases, pore size in gel matrix ___

A

decreases

27
Q

What are the different topologies (conformations) of DNA molecules?

A

Linear, relaced circular, and supercoiled

28
Q

T.F. The topology of DNA strands affect their rate of migration in gels?

A

True

29
Q

In cells what is the charge of supercoiled DNA?

A

negative

30
Q

What can speed up the migration rate of DNA fragments during agarose gel-electrophoresis?

A

greater voltage

31
Q

What are some factors that DO NOT influence the rate of migration of DNA molecule’s during agarose gel-electrophoresis?

A

The %GC content or sequence of a DNA molecule

32
Q

Requirements for DNA synthesis in vitro?

A

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
Q

What direction does DNA synthesis always proceed?

A

3’

34
Q
A
35
Q

What does PCR stand for?

A

polymerase chain reaction

36
Q

Mullis’ insight (associated with PCR)

A

enzymatic copying of double-stranded DNA using 2 primers, complementary to opposite strands could lead to exponential increase in amount of target sequence

37
Q

PCR required DNA to be cycled repeatedly through how many temperatures?

A

3

38
Q

Generally how many cycles of PCR?

A

30-35 (in theory more than a billion-fold amplification)

39
Q

Order of PCR steps

A

1) Denaturation
2) Annealing
3) Elongation or extension

40
Q

Denaturation PCR step

A

94-96 C, dsDNA denatured to ssDNA

41
Q

Annealing PCR step

A

50-65 C, primers bind to their complementary sequences, Tm is dependent on length and base composition of primers

42
Q

Elongation or extension PCR step

A

72 C, DNA polymerase binds to the annealed primers and extends DNA at the 3’ end of the chain

43
Q

What are the ‘ingredients” in PCR?

A

dinucleotide triphosphates, Mg2+, primers, template DNA, thermostable DNA polymerase, a salt, Tris (pH control), plus stabilizers

44
Q

taq

A

Thermus aquaticus, a thermostable DNA polymerase

45
Q

What are Dinucleoside triphosphates (dNTPs)

A

dATP, dCTP, dGTP, dTTP

46
Q

What are primers in PCR?

A

short molecules of ssDNA (aka oligonucleotides),

47
Q

What is the results of priming between two logos annealed to opposite strands?

A

gives exponential growth of product

48
Q

What effects the size of the PCR products?

A

How far apart the annealing sites of the 2 primers

49
Q

How long are primers typically?

A

18-25 bp (enough to have binding sites, but also not so long they cost a lot of energy)

50
Q

Applications of PCR?

A

Amplifying target sequences for further study, detection of rare DNA sequences,

51
Q

What is PCR not good at?

A

Determining abundance of the rare sequences

52
Q

What type so growth does the early phases of PCR experientces?

A

exponetial

53
Q

What type of growth does later cycles of PCR experience?

A

linear

54
Q

What types of growth does end stage of PCR experience?

A

plateau

55
Q

What phase provides the best information to estimate starting amount of DNA (or RNA) template in PCR?

A

Log-linear phase

56
Q

Alternative name for real-time PCR

A

quantitative PCR (qPCR)

57
Q

qPCR procedure

A

uses a reporter dye to monitor the PCR product

58
Q

What dye is used most commonly in qPCR?

A

SYBR green

59
Q

What grooves does SYBR fluorescence most commonly bind to ?

A

minor groove in dsDNA

60
Q

Application of qPCR?

A

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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