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Pure culture
ASEPTIC TECHNIQUE
Sterilizing Instruments
Bunsen Burner Flame
Aseptic Transfer
Removing Cap
Flaming Tube
Holding Tube
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Hand Mixing
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Plate inoculation
Invert Plate
Streaking Plate
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Video:
Aseptic technique
Streak plate
1. Inoculate
2. Incubate
3. Isolation
4. Inspection
5. Information gathering
6. Identification
1. Inoculation
Introduction of a small sample inoculum
into a container of media to produce a
culture of observable growth
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Culturing Microorganisms
Obtaining Pure Cultures
Cultures composed of cells arising from a single
progenitor
Progenitor is termed a CFU
Streaking Plate
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2. Incubation
Temperature-controlled chamber
Microbe multiplies and produces
macroscopically observable growth
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3. Isolation
If an individual bacterial cell is separated
from other cells and has space on a nutrient
surface, it will grow into a mound of cells-a colony.
A colony consists of one species.
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4. Inspection
Inspection observation; macroscopic and microscopic
Pure culture grows only single known species of
microorganisms
Mixed cultures hold two or more identified species or
microorganisms
Contaminated culture once pure or mixed culture that
has unwanted microbes growing
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5. Information gathering
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6. Identification
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growth inhibitors
liquid/solid
temperature
source of energy
sources of carbon, nitrogen, ...
Aseptic technique:
Necessary equipment
Procedure
1.
2.
3.
4.
5.
6.
With the loop, spread the inoculum back and forth across the upper 1/4 of the
plate, keeping the lines of inoculation very close together (area 1 in figure
below).
Isolated colonies are not expected in this area. Do not use strong pressure,
which will break the surface of the agar. Use the end of the loop, not its side
when streaking. Dispose of the loop in the biohazard bucket on the bench.
Turn plate approximately 90oC. Streak the plate as indicated in the figure
(area 2) across about 1/4 of the plate. Dispose of the loop.
Repeat step 2 one or two times more.
In area 3 and/or 4 single colonies should appear.
Label plates on the bottom and incubate inverted at 37oC.
Streaking
Remember
1.
2.
3.
4.
5.
6.
7.
Bacteria
Are everywhere!
On every surface of the body
Including digestive tract
Harmless
Beneficial
Pathogenic
Absorb nutrients and release toxins that damage cells
and tissues.
8. Bacterial toxins can cause disease even when
bacteria are destroyed
9. Bacteria are Prokaryotes