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MCB 101- LEC > Lecture 4: Culture Preservation and Maintenance_1 > Flashcards

Lecture 4: Culture Preservation and Maintenance_1 Flashcards

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1
Q

Goal of culture preservation (2)

A
  1. Retain viability and functionality of the stock culture for a long period of time
  2. Maintain purity and trait of being “true-to-type”
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2
Q

_____ means to maintain the live form or viability of the organism

A

Viability

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3
Q

______ is maintaining the normal characteristics or phenotypes of the organisms

A

Functionality

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4
Q

Importance of culture preservation (4)

A
  1. ensures the availability, supply and functionality of microbial strains for future studies
  2. as reference for standardized assays and tests
  3. for taxonomic purposes
  4. as valuable stock for biotechnological applications
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5
Q

Important principles for preservation and maintenance of stock cultures (3)

A
  1. Reduction in the temp of growth of the organism
  2. Dehydration or desiccation of the medium of growth
  3. Limitation of nutrients to the organism
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6
Q

Important things to remember in culture preservation (7)

A
  1. Zero tolerance to contaminants
  2. Number of replicates
  3. Competent personnel
  4. Knowledge about the culture
  5. Proper labeling for containers
  6. Proper protective gear/clothing
  7. Proper and secure inventory
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7
Q

Considerations of periodic transfer to fresh media (3)

A
  1. Time interval pf transfers
  2. Proper medium
  3. Proper storage temperature
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8
Q

Name the 7 culture preservation methods

A
  1. Storage in distilled water
  2. Drying
  3. Lyophilization
  4. Periodic transfer to fresh media
  5. Cryopreservation
  6. Overlaying cultures with mineral oil
  7. Ready to use preservation kit
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9
Q

Aerobic organisms are usually inoculated in

A

Agar slants and petri plates

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10
Q

Anaerobic organisms are usually inoculated in

A

Agar stabs

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11
Q

Advantages of periodic transfer to fresh media (2)

A
  1. Easy to prepare
  2. Does not require expensive equipment
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12
Q

Disadvantages pf periodic transfer to fresh media (3)

A
  1. Risk of spontaneous mutation
  2. Prone to contamination
  3. Requires larger space to maintain several cultures
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13
Q

The aim of overlaying cultures in mineral oil (2)

A
  1. Limit the availability of O2
  2. Reduce the rate of dehydration of medium
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14
Q

overlaying cultures in mineral oil is most effective to _______ bacteria

A

Aerobic

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15
Q

Advantages of overlaying cultures in mineral oil (3)

A
  1. Cheap and readily available
  2. Easy to prepare
  3. Working culture can be directly obtained from mineral oil stock
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16
Q

Disadvantages of overlaying cultures in mineral oil (2)

A
  1. Requires larger space to maintain several cultures
  2. Viability of microorganisms varies with species
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17
Q

Microorganisms which can degrade mineral oil (3)

A
  1. Pseudomonas putida
  2. Rhodococcus erythropolis
  3. Bacillus thermoleovorans
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18
Q

Freeze drying (lyophilization) employs _____ of ice or other frozen solvents from the material through the process of ______ and the removal of bound water molecules through the process of ____

A

removal, sublimation, desorption

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19
Q

Examples of cryoprotective agents (lyoprotectants)

A

Skim milk, glycerol etc

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20
Q

The efficacy period of maintaining the cultures subjected to freeze drying

A

>20 years

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21
Q

Is considered as one of the most effective method in culture preservations

A

Freeze drying

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22
Q

During freeze drying, what will happen to the cultures after the removal of water

A

Cells in a inactive state

23
Q

Advantages of freeze-drying

A
  1. Easy to replicate
  2. Light and easy to transport
  3. Less opportunity for changes in characteristics of culture
24
Q

What glassware does the lyophilized cells are kept?

A

Borosilicate glass

25
Q

Examples of freeze dryers

A
  1. Typical manifold dryer
  2. Shelf freeze dryer
  3. Combination lab dryer
  4. Console manifold freeze dryer
26
Q

How small is the borosilicate vial containing the lyophilized culture?

A

smaller than a pen

27
Q

What storage temperature is ideal for preservation in agar ?

A

Ordinary refrigeration (4-10 degree Celsius)

28
Q

What storage temperature is ideal for cryopreservation?

A
  • 192 degree Celsius
29
Q

Cryopreservation employs the use of _____ and ______ . Provide also its purpose

A

liquid nitrogen - to preserve the culture at - 192 degree Celsius, glycerol - to prevent lethal damages to the cells

30
Q

Commonly used cryoprotectants in cryopreservation (2) provide percent concentrations

A

Dimethyl sulfoxide (DMSO 5-10%) and glycerol (5-20%)

31
Q

Purpose of cryoprotectants in cryopreservation (3)

A
  1. Aid in preserving cells
  2. Encourage dehydration
  3. Minimize solution effects
32
Q

Properties of good cryoprotectants in cryopreservation (4)

A
  1. Non-toxic
  2. Not biodegradable by the organism to be preserved
  3. stable and compatible to target microorganism
  4. readily available
33
Q

cryoprotectant/s for animal cells with concentration

A

DMSO (5-10%) or glycerol (5-10%)

34
Q

cryoprotectant/s for bacteria with concentration

A

glycerol (5-10%)

35
Q

cryoprotectant/s for yeast with concentration

A

glycerol (10%)

36
Q

cryoprotectant/s for protozoa with concentration

A

DMSO (5-10%) or glycerol (10-20%)

37
Q

cryoprotectant/s for plant cells with concentration

A

DMSO (5-10%) or glycerol (5-10%)

38
Q

cryoprotectant/s for animal viruses (free) with concentration

A

none

39
Q

cryoprotectant/s for animal viruses (infected cells) with concentration

A

DMSO (7%)

40
Q

What is the drying temperature

A

45 degree Celsius

41
Q

drying preservation method is limited to what type of organisms?

A

spore and cyst formers (dormant structures)

42
Q

What structures are being dehydrated during drying preservation?

A

Vegetative cells and dormant structures

43
Q

Drying preservation is applicable to ____ organisms especially those who produces _____

A

soil, endospores

44
Q

In drying preservation where does the suspension of pure cultures inoculated?

A

sterile paper disc

45
Q

aside from paper disc, what else is used in drying preservation method?

A

porcelain beads

46
Q

sterile distilled water is more effective in preserving what cultures?

A

spores or conidia of molds

47
Q

How does mold spores harvested before preserving in sterile distilled water?

A

using moistened swab stick

48
Q

storage temperature for culture preservation in sterile distilled water

A

room temperature

49
Q

example of ready-to-use preservation kit for culture preservation methods

A

“protect” of ThermoFisher Scientific

50
Q

How do we revive the lyophilized and cryopreserved cultures?

A

rehydrated of lyophilized cultures and thawing of frozen cultures and inoculated on fresh media

51
Q

3 ways in checking purity of preserved microorganisms

A
  1. Evaluating the cultural characteristics
  2. Gram staining and microscopy
  3. performing 16S rRNA gene sequencing
52
Q

How do we revive the preserved stock from agar, mineral oil and sterile water?

A

streak for isolation on an appropriate medium

53
Q

These are organizations that maintain authentic pure culture of microorganism

A

culture collection centers

54
Q

Example of institutions with standing name in nomenclature of prokaryotes

A
  1. American type culture collection (ATCC)
  2. National Collection of Type Cultures (NCTC)
  3. Japan Collection of Microorganisms (JCM)
  4. Philippine National Collection of Cultures (PNCC)

MCB 101- LEC (6 decks)

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