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MCBM- Exam 1 > DNA Replication > Flashcards

DNA Replication Flashcards

1
Q

Southern Hybridization

A

Hybridization of a probe fragement to a DNA target

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2
Q

Northern Hybridization

A

Hybridization of a probe fragement to a RNA target

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3
Q

Requirements of PCR

A

Template (ss or ds)

Pair of primers

4 deoxyribonuclotides

Taq DNA polymerase

Appropriate buffer

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4
Q

PCR: DNA is denatured at __ degrees and strand extension occurs at __ degrees

A

Denatured: 95 degrees celcius

Strand extension occurs at 72 degrees celcius which is the optimal temperature of Taq

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5
Q

Equation for PCR

A

2^n where n = number of cycles

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6
Q

How does microarray/chip based hybridization work?

A

Large numbers of gene-specific oligonuclotides printed/spotted on a chip are hybridized to total RNA and the number and intensity of hybridizing spots are determined

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7
Q

What is the medical function of microarray/chip based hybridization

A

Allows us to address a treatment-specific question: Is there a difference in gene expression pattern between responders and non-responders of drugs

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8
Q

Medically important applicaitons of DNA

A

Disease diagnosis

Prenatal diagnosis

Bacterial infections

Virus infection and viral load

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9
Q

DNA matches are obtained by experimental analysis needs to be confirmed by

A

Statistical analysis

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10
Q

Forensic medicine relies on

A

Variable number of tandem repeats (VNTR)

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11
Q

Cell doctrine

A

Every cell in the body arises from an existing cell

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12
Q

In what phase does DNA replication occur?

A

S phase

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13
Q

2 main phases of cell cycle

A

Interphase + Mitosis

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14
Q

Interphase consists of

A

G1

S

G2

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15
Q

Direction DNA synthesized in

A

5’ to 3’

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16
Q

5’ end of DNA contains

3’ end of DNA contains

A

5’: phosphate

3’: hydroxyl

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17
Q

E. coli polymerase types

A

DNA Pol 1-3

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18
Q

Processivity of DNA Pol I-III

A

DNA Pol I: low processivity

DNA Pol II: higher processivity

DNA Pol III: highest processivity

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19
Q

Which DNA Pol do DNA repair

A

Pol I and II

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20
Q

What is the function of DNA Pol III

A

DNA replication

5’ –> 3’ polymerization

3’ –> 5’ exonuclease

21
Q

What is the function of DNA Pol I

A

5’ –> 3’ polymerization

5’ –> 3’ exonuclease

3’ –> 5’ exonuclease

22
Q

dnaA =

A

initiator protein

23
Q

dnaB =

A

helicase

24
Q

dnaG =

A

primase

25
Q

In E. Coli replication, the initiator protein binds to the origin and denatures ___ base pairs

A

A-T

26
Q

dsDNA is unwound by

A

Helicase (dnaB) in an ATP-dependent manner

27
Q

Unwound strands of DNA are stabilized by

A

ss binding protein

28
Q

What relieves overwinding of of DNA as one part unwinds

A

Type II topoisomerase (gyrase)

29
Q

Primase synthesizes

A

15 nt long RNA primers

30
Q

What is the absolute requirement for RNA replication

A

Primers

31
Q

DNA polymerase requires ___ and __ for synthesis of a polynucleotide strand

A

Template

Free 3’ OH group

32
Q

DNA polymerase I- function

A

Removes RNA primers which are then replated by extending DNA synthesis

33
Q

E. Coli polymerase III is a __ enzyme

A

Holoenzyme

34
Q

What are the subunits of E. Coli Polymerase III (holoenzyme)

A

Catalytic core of 3 subunits (alpha-epsilon-theta)

Clamp subunits (beta, beta) for processivity

Clamp loader (gamma-complex)

Assembly subunit (tau, tau)

35
Q

Logical target for inhibiting bacteria replicaiton

A

Enzymes of DNA replication

36
Q

Streptomyces spheroides and streptomyces niveus produces

A

Novobiocin

37
Q

What is novobiocin

A

Replicaiton/transcription inhibitor

38
Q

Apart from novobiocin, what are some other synthetic antibiotics

A

Fluoroquinolones = levofloxacin and ciprofloxacin

39
Q

Type I vs. Type II topoisomerase

A

Type I makes a cut on only 1 strand of DNA

Type II = gyrase and makes 2 cuts (one on each strand of DNA)

40
Q

What is the target enzyme for fluoroquinoles?

A

Bacterial gyrase

41
Q

Gyrase is a ___ (structure)

A

Heterotetramer (GyrA2GyrB2)

42
Q

How does gyrase cleave DNA?

A

Tyr122 on GyrA forms covalent linkage with the 5’P of DNA

GyrB carries out ATP hydrolysis

43
Q

How do fluoroquinolones inhibit Gyrase?

A

They prevent the revesible ligation step carried out by GyrA

44
Q

What competes with ATP for binding to GyrB

A

Novobiocin

45
Q

What is the target enzyme for anticancer drugs?

A

Mammalian Topo I

46
Q

Topoisomerase I binds to

A

3’ end of phosphodiester back bone

47
Q

How does Topotecan work?

A

It stabilizes topoisomerase I-DNA complex and prevents the religation step = inhibition of replicaiton

48
Q

Resistance to quinolone may be caused by point mutations to

A

N-terminal end of GyrA (most mutations)

C-terminal end of GyrB (few mutations)

MCBM- Exam 1 (7 decks)

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