LAB 3: PREPARATION AND STERILIZATION OF CULTURE MEDIA
SCHOOL OF TECHNOLOGY
INDUSTRY
UNIVERSITY SCIENCES
MALAYSIA
DEGREE IN BIOPROCESS
IBG 102
BIOLOGY FOR
TECHNOLOGIST
Name
|
1.
LOH
SHI WEI (137602)
2.
LAI
CHONG SING (137592)
3.
SITI
NORASYIKIN BINTI SALMI (137673)
4. SITI
NUR SUHAILI AFIQAH BINTI SARIMAN (137674)
5.
NUR
LIYANA ATHILAH BINTI MOHD AFFANDI (137636)
|
Title
|
LAB 3: PREPARATION AND STERILIZATION
OF CULTURE MEDIA
|
Date
of practical
|
25/09/17
|
Date of Report Submitted
|
01/10/17
|
Lecturer
|
DR. TYE
|
LAB 3: PREPARATION AND
STERILIZATION OF CULTURE MEDIA
Culture
medium is a liquid or gel designed to support growth of microorganism or cell.
There are different type of media for growth different types of cells.
The
most common growth media for microorganism are nutrient broth or LB medium
(Lysogeny Broth). Liquid media are often mixed with agar and poured into petri
dishes to solidify. These agar plates provide a solid medium on which
microorganism can culture. The microorganism will grow on the surface of the
agar and this make examination work so much easier as the colonies of
microorganism remain stationary and clearly visible.
The composition of
growth media is listed as below:
3.0 g/L "Lab-lemco" powder (a beef extract).
2.0 g/L yeast extract.
5.0 g/L peptone (a nitrogen source)
5.0 g/L sodium chloride
8.0 g/L agar powder
3.0 g/L "Lab-lemco" powder (a beef extract).
2.0 g/L yeast extract.
5.0 g/L peptone (a nitrogen source)
5.0 g/L sodium chloride
8.0 g/L agar powder
Autoclaving:
Autoclaving is the method that can established for a variety of application especially an apparatus (as for sterilizing) using this steam under high pressure (121 °C) for 15 minutes.
Autoclaving is the method that can established for a variety of application especially an apparatus (as for sterilizing) using this steam under high pressure (121 °C) for 15 minutes.
An
autoclave is in essence a large pressure cooker; a chamber which may
sealed off against surrounding microorganism. Materials for sterilization are
placed in the chamber. The incoming steam displace cooler air through an
exhaust valve; this valve closes when the cell cooler air has been vented.
OBECTIVE
To prepare sterile nutrient agar for culturing
microorganisms
MATERIALS
AND REAGENT
Commercial nutrient agar
Balance
Distilled water
Scott bottles
PROCEDURE
1. Self Made Nutrient Agar
Preparation
1. 1.5g/L
‘ Lab-lemco” powder, 1.0g/L yeast extract, 2.5g/L peptone, 2.5g/L sodium
chloride and 7.0g/L agar powder were weighed and put into the two 500ml of
Scott bottles each respectively.
2. 500
ml of distilled water was measured using measuring cylinder and poured into the
Scott bottles each respectively.
3. The
mixture inside the Scott bottles was shake until it dissolved completely with
the distilled water.
4. The
Scott bottles were labeled with the marker pen.
5. The
Scott bottle were loosely recapped and autoclaved at 121°C/15psi for 15
minutes.
6. After
autoclaving, The Lb agar was removed from the autoclave and was allowed to cool
before tighten the cap of the Scott bottle.
2. Commercial Nutrient Agar
Preparation
1. 50g
of commercial nutrient agar was measured and put into the two Scott bottles
each respectively.
2. 500ml
of distilled water was measured and poured into the two Scott bottles each
respectively.
3. The
mixture in the Scott bottles was shake until the powder dissolved completely in
the distilled water.
4. The
Scott bottles were labelled with the marker pen
5. The
Scott bottles were loosely recapped and autoclaved at 121°C/15psi for 15
minutes.
6. After
autoclaving, the media was removed and was allowed to cool before tighten the
cap of the Scott bottle.
RESULT
DISCUSSION
The preparation of culture media which contains
nutrients needed by microorganisms is needed for cultivation of microorganisms.
Culture media can be prepared manually or commercially. Preparing culture media
manually can be labour intensive and very time-consuming, but appropriate
amount of media could be prepared without excess waste. Commercial culture
media offers a time-saving solution, but it does not address the need to
accurately predict the quantities required for future testing. There are
several precautions that we need to take during the experiment.
Firstly, we must read the label and instruction on the container before
use during preparation of commercial media, BHI media and TSAYE media. In the
progress of experiment, use distilled water to clean all the apparatus.
Measuring cylinder is used to measure the volume of distilled water required
accurately. Besides, all the Scott bottles must be labelled first. The proper
receiver for the material must be selected. The receiver’s weight plus the weight
to be measured must not exceed the maximum load for the balance. The size and
shape of the receiver should permit it to fit into the space and on the balance
pan without interfering with any operation. It is important that the receiver
is clean and in dry condition. The surrounding of the pan and the pan of the
balance is made sure to be clean. The receiver is placed in the center of the
pan of the balance. Then, the tare key is pressed every time after the receiver
with substances is put onto the balance to avoid zero error on the balance. The
powdered material is carefully put into the Scott bottles using a spatula until
the desired amount is added. Handle with care to avoid spilling. If solids are
spilled, remove the receiver and sweep out all the spilled material from the
balance using a brush. The spilled material must be properly
disposed.
When dissolving the agar powder, the distilled water
in the measuring cylinder should not be poured all out into the beaker, because
there should be some distilled water reserved for the washing of leftover
powders from the weighing container into the beaker. This is done to ensure
that all the agar powder has dissolved in the culture medium. The media should
be stirred evenly by spatula before pouring into the Scott bottles to ensure
the powders mix well with distilled water. Agar plates provides a solid medium
where microbes cultured. Bacteria hardly decompose the agar as they are solid.
Autoclaving is a method of sterilization, high
temperatures is achieved by steam under pressure in an autoclave. Before
autoclaving, the caps of the Scott bottles should be loosened. This is because
the autoclave machine operates under high steam pressure and by loosening the
cap will allow expansion of the bottle so that the bottles will not be broken.
The autoclaving machine should be closed tightly so that the media can be
sterilized at 121 °C. The temperature is checked so it is always
maintained at 121 ᵒC and with steam is continuously forced into the
chamber, the pressure is ensured to reach 103 kPa above the atmospheric
pressure. The time for destruction of the most resistant bacterial spore is
about 15 minutes. Then, the exhaust valve is opened to ensure the pressure
drops to atmospheric pressure before removing the Scott bottles from the
autoclave chamber. The cap of the Scott bottles that are removed from the
machine should be tightened. The media are cooled down.
CONCLUSION
In conclusion, the objective of the experiment which is to prepare sterile nutrient agar for culturing microorganisms is success.
REFERENCES
https://www.sciencebuddies.org/science-fair-projects/references/grow-microbes-agarIn conclusion, the objective of the experiment which is to prepare sterile nutrient agar for culturing microorganisms is success.
REFERENCES
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