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MICB 3301 Lab > Quiz #5: 11B, 12, 13, 14, and 15 > Flashcards

Quiz #5: 11B, 12, 13, 14, and 15 Flashcards

1
Q

Lifecycle of helper T cell recognition

A

Th cell recognizes antigen bound to MHC class II of a B cell or macrophage

Th cell produces cytokines that direct further immune response

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2
Q

What was KZH in the T cell experiment?

A

KZH was a T helper hybridoma, created by fusing normal helper T cells from mice immunized with antigen lysozyme + a T lymphoma cell with a reporter

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3
Q

In T cell experiment, what reporter does KZH have and how does it work?

A 
Gene lacZ (makes B-galactosidase) connected to IL-2 gene (makes cytokine IL-2) which is activated upon antigen binding. 
Cell is lysed to release B-gal using TritonX-100
B-gal now can now hydrolyze CPRG to cause color change
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4
Q

In T cell experiment, what was the antigen and the antigen presenting cell?

A

Antigen is lysozyme

Antigen presenting was mouse B cell hybridoma LK35.2

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5
Q

In T cell experiement, why did we culture the cells in 5% CO2

A

Because it established equilibrium with bicarbonate in growth medium, forming a buffer that maintains pH at about 7.4

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6
Q

In T cell exp, what ingredients were in RPMI-1640?

A 
L-glutamine, 
pyruvate, 
2-mercaptoethanol, 
HEPES, 
fetal bovine serum, 
antibiotics gentamicin & penicillin & streptomycin
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7
Q

What temperature was T cell experiment plate incubated at day 2?

A

37ºC

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8
Q

Narrow-spectrum antibiotics

A

effective only against a limited number of pathogens

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9
Q

Broad-spectrum antibiotics

A

attack many different kinds of bacteria

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10
Q

Cidal antibiotics

A

kill microorganisms

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11
Q

Static antibiotics

A

Do not kill but inhibit growth

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12
Q

Penicillin

A

First medically useful antibiotic

Produced by Penicillium notatum and Penicillium chrysogenum

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13
Q

Streptomyces

A

Genus of Actinomycetes

Includes antibiotics: 
Streptomycin
Kanamycin
Vancomycin
Rifampin
Chloromycetin
Tetracycline
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14
Q

What is an antibiotic that is now produced synthetically?

A

Chloromycetin is now chloramphenicol

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15
Q

Applications of antibiotics

A

veterinary medicine, animal feed supplementation, plant disease control, food preservation

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16
Q

Provide generic name, biological source, and MOA for beta-lactams

A

Penicillins and cephalosporins

Penicillium and cephalosporium

Inhibits synthesis of bacterial cell wall peptidoglycan

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17
Q

Provide generic name, biological source, and MOA for macrolides

A

Erythromycin

Streptomyces erythreus

Inhibits rRNA associated with 50s LSU ribosome

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18
Q

Provide generic name, biological source, and MOA for aminoglycosides

A

Streptomycin, neomycin, gentamicin

S. griseus, S. fradiae, Micromonospora purpurea

Inhibits 30s SSU ribosome

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19
Q

Provide generic name, biological source, and MOA for tetracyclines

A

Tetracycline

Streptomyces aureofaciens

Inhibits binding of aminoacyl-tRNAs to ribosome

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20
Q

Which antibiotic producing microbes did we use in 12A?

A

Penicillium notatum and Streptomyces griseus

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21
Q

Which bacteria did we test in 12A and their reaction to each antibiotic?

A

E.coli: none P, some S
B.subtilis: full P, none S
S.aureus: full P, some S
Micrococcus luteus: full P, full S

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22
Q

What temp were plates incubated at for 12A after days 1 and 2

A

Day 1- 30ºC

Day 2-37ºC

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23
Q

What is the Kirby-Bauer method?

A

Inoculated Mueller-Hinton or TSA plates with antibiotic discs, measure inhibition zones

24
Q

Minimal inhibitory concentration (MIC)

A

The lowest concentration of antibiotic that will prevent growth, proportional to the diameter of the inhibition zone around the disc

25
Q

What method was used to standardize the disc diffusion bacterial inoculum turbidity and what value was used?

A

McFarland standards, 0.5 used

26
Q

What antibiotic discs were used in 12B?

A

Erythromycin (E15)
Penicillin (P10)
Tetracycline (TE30)
Gentamicin (GM10)

27
Q

What bacteria were used in 12B?

A

E.coli and S.aureus at McFarland 0.5

28
Q

What stage of the bacterial growth curve is the bacteria most susceptible to antibiotics?

A

Log/exponential phase

29
Q

What are the intermediate values (in mm) of each antibiotic— sensitive is anything more and resistant is anything less

A

E15: 14-17
P10: 12-21 (21-28 for staphylococci)
T30: 15-18
G10: 13-14

30
Q

Horizontal gene transfer

A

Process where genes are transferred from one mature independent organism to another

31
Q

Conjugation

A

The process of genetic transfer that involves cell-to-cel contact between donor and recipient cells

32
Q

Transformation

A

free DNA is taken up by a cell and integrated into its genome

33
Q

Transduction

A

Transfer of genes between bacterial or archaeal cells by bacteriophages

34
Q

Competent cells

A

Able to take up free DNA

35
Q

What streptomycin resistant strain of bacteria was used in Ex. 13

A

Acinetobacter calcoaceticus

36
Q

Ex. 13 incubation temperature

A

30ºC

37
Q

Which of the 4 sectors on the plate should have no growth and why?

A

DNA only because there are no cells present

38
Q

What temperature was the DNA incubated at in Ex. 13 and why

A

60ºC to lyse cells and have purified DNA

39
Q

R factors

A

Conjugative plasmids the carry genes for resistance to antibiotics

40
Q

tra genes

A

Donor genes that encode proteins that help carry out conjugation

41
Q

F (sex) pilus

A

tra gene product that mediates contact between donor and recipient for conjugation

42
Q

In Ex. 14, what plasmid was being transferred? What are some key features of this plasmid?

A 
pBBR1MCS-2: 
Kanamycin r gene 
lacZ gene
mob 
multiple cloning site (MCS)
43
Q

Restriction endonuclease

A

enzymes that cut DNA into pieces at the restriction sites (in MCS region)

44
Q

What is the donor E.coli strain used in Ex. 14, what antibiotics is it sensitive/resistant to

A

Strain S17-1 containing tra genes for plasmid transfer.

Sensitive to nalidixic acid, resistance to kanamycin

45
Q

What is the recipient E.coli strain used in Ex. 14, what antibiotics is it sensitive/resistant to

A

Strain DH5å

Sensitive to kanamycin, resistant to nalidixic acid

46
Q

How did the three plates grow in exercise 14?

A

Nalidixic acid: recipient only, white
Kanamycin: donor only, blue
Nal+Kana: mate growth, blue

47
Q

Bacteriophages or phages

A

viruses that infect bacteria, basically a fragment of genetic material with protein coating

48
Q

Bacteriophage infection cycle

A

Phage binds via tail fibers to a cell receptor
Phage genetic material, stored in capsid, is injected into host
In host, gentic material either is used to make more phages and lyse cell (lytic) or forms prophage in genome and becomes latent (lysogenic)

49
Q

Phage titer

A

The concentration of infectious phage particles per mL of growth medium

50
Q

Plaques

A

clear zones in a lawn of bacteria where phages killed cells

51
Q

What broth were E.coli in in Ex 15?

A

Luria-Bertani (LB) broth

52
Q

What facilitates phage attachment to bacterial receptors?

A

magnesium ions

53
Q

Temperate bacteriophages and example

A

carried by host cells for generations– lysogenic

Lambda temperate bacteriophage infects E. coli, can enter lytic or lysogenic cycle tho

54
Q

Repressor protein cI

A

product of phage gene that regulates the lysogenic cycle of lambda phage, binds to operator region

55
Q

Regulatory protein Cro

A

promotes lytic cycle in lambda phage, binds to same operator as cI but opposite effects

MICB 3301 Lab (2 decks)

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