Microalgae culture are essential
nowadays when conducting the research in aquaculture production, specifically
to mollusc culture and other types of cultures. Microalgae culture may be
‘unialgal’, which means they contain only one kind of microalgae, usually a
colonial population, but somehow may contain bacteria, fungus or protozoa.
Microalgae culture also may be ‘axenic’, meaning that they contain only one
microalgae without bacteria, fungus or protozoa. There are several techniques
or methods that can be used in isolation of microalgae, which are single cell
picking, streaking method and serial dilution method.
Single cell picking is the best method
which can be used in order to get axenic cultures. This is due to the this
method is emphasise to get only one cell by using micropipette. Although this
technique is quite tough to do, but the result from this technique is very
amusing because there are no bacteria or any other fungus that can be found
when culturing the microalgae.
Streaking method and serial dilution
method is more profoundly to get the unialgal cultures. This is because this
method does not cover only one single cell or one single species, but this
method will cover all the microalgae that the sample solution have. In other
words, there are possibilities that this culture may have contained the
bacteria or fungus at the end of this experiment.
OBJECTIVE
To practices on different microalgae
isolation techniques
METHODOLOGY
Single cell picking
I.
Pasteur pipette is held in the hottest
region of the flame, supported on left hand by a hand and on the right by
forceps.
II.
When the glass is soft, the Pasteur
pipette is quickly removed from the flame with a gentle pull to produce thin
tube.
III.
The forceps is then relocated to the
appropriate region in the thin tube.
IV.
The forceps is used to gently bend the
thin area so that it breaks, which will form micropipette.
V.
From the sample solution, it is placed
on the glass slide one drop and observed under the microscope.
VI.
If the cell is too abundant in that
drop, the one drop from sample solution is diluted by adding one drop of
seawater.
VII.
Step V and Step VI is repeated to get
one single species on the glass slide.
VIII. The single cell is being picked from the
sample solution that have been diluted by using micropipette.
I.
The loop is flamed and allowed to cool.
II.
The cap of test tube contained sample
solution is removed and being flamed at the neck of the test tube.
III.
The loop is inserted into the test tube
and being dipped into the sample solution, then withdrew it.
IV.
Again, the neck of the test tube is
being flamed.
V.
A loopful of the culture is placed on
the agar medium on the area 1.
VI.
The flop is flamed and cooled about 5
seconds by touching an unused part of the agar medium surface close to the
periphery of the plate, and then it is dragged rapidly several times across
surface area 1.
VII.
The loop is removed and the petri dish
contained the medium agar is closed.
VIII.
The step is repeated by the loop is
reflamed and cooled.
IX.
The petri dish is turned 90o anticlockwise
then the loop is touched to a corner of the medium culture in area
1 and it is dragged several times across the agar medium in area
2.
X.
Step VII until Step IX are repeated
until there are 3 area in the agar medium which have been dragged by the loop.
XI.
Incubate the agar medium under the
light.
I.
1ml of sample solution
is took and poured into test test tube.
II.
9ml of seawater is
added into the test tube for dilution purpose.
III.
This test tube is
labelled as 10-1.
IV.
Then 1ml of solution in
test tube 10-1 is took and poured into the new test tube, which
then 9ml of seawater is added into that test tube by labelled as 10-2.
V.
Steps are repeated
until there are 10 test tubes which last test tube is labelled as
10-10.
VI.
The test tubes is
incubated under the light.
RESULTS
Single cell picking
Figure 1: Sample solution before being diluted
Figure 2: After diluted several times
Streaking method
Figure 3: After 4 weeks of culture
DISCUSSION
Based from the results that showed in each method,
it can says that each method is suitable for us to practice those methods.
Although there are several methods that were not very successful in isolate the
microalgae, like streaking method, but it shown that there are several human
error that lead to that. For example, in streaking method, it shown that there
were no microalgae that can be cultured at the end of the experiment. Actually
this may be causes by several factors during handling the experiment, such as
the wire loop that have been used during this method is not fully sterilized
and may contaminated which will lead to the failure in culture the microalgae.
On top of that, during incubation period, we can see that the place that we
incubated the microalgae was quite crowded which may lead to the insufficient of
light that causes the growth of microalgae being stunt. As the solution for
those problems, what we can do in order to improve the result of this
experiment is that by ensuring all the materials that are going to be use in
the experiment are fully sterilized. In addition, during handling the wire
loop, please make sure that the wire loop is not touched any other things that
may causes contamination. Lastly, the place that we are going to incubate or
isolate the microalgae must suitable and have sufficient light to ensure the
microalgae grow.
In single cell picking method, the purpose why the
forceps is gently bend the thin micropipette is to ensure the tips of the tube
smooth, so that the cell is not break during the picking of cell happen. The
broken tips of tubes may lead to burst of the cell, which can causes the
failure in isolate the microalgae.
CONCLUSION
Based from the discussion above, this experiment
shown that all the methods in isolate the microalgae can be used. Single cell
picking is most profoundly the most suitable method to get the axenic culture,
while the rest are more tend to get the colonial microalgae.
REFERENCES
Parvin, M., Zannat, M. N., & Habib, M. A. B. (2007). Two Important Techniques for Isolation of Microalgae. Asian Fisheries Science, 20(1/2), 117.
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