Marine Biotechnology Lab Collection and handling samples for Algal isolation

Activity 1 Students have to complete the preparation of sock solutions of the F/2 medium. Adjust the salinity of seawater to 25‰ by using a refractometer and filter 2 liter of the seawater through double layer of Whatman filter paper. Students have to prepare 2 flasks each of 2 liter capacity and containing 950 ml of seawater to which 1 ml of NaNO3 solution was added. To avoid precipitation upon autoclaving dissolve 0.4g/l of trihydoxymethyl aminomethane in 50 ml of dist water and add to flask.

Autoclave 100 ml of 0.1 N NaOH and 0.1 N HCl. Autoclave 3- 250 ml Erlenmeyer flasks and 20 screw –capped test tubes. After autoclaving and cooling down the seawater, aseptically add the sterilized nutrients, trace metals and vitamin solutions while mixing. To reduce the risk of contamination, all work should be done in biological safety cabinet.

Adjust the pH of flask to 8 – 8 Adjust the pH of flask to 8 – 8.5, determine the volume of base or acid added. Dispense the sterilized medium from the flask into the 3- 250 ml Erlenmeyer flasks (100-150 ml each) and the 15 screw –capped test tubes (exactly 9ml each).

Collection and handling samples for Algal isolation Activity 2 Students are requested to bring the following samples A 2-l bottle or jar filled with seawater. A cup filled with small amount of bottom sediment. Some dead leaves, algae, or any biological material found floating on the sea surface or on the seashore.

In the lab... In the laboratory, students have to do the following. Vigorously shake the water sample and filter about 100 ml through 0.45 μm Millipore filter paper. Transfer the filter paper into one of the 250 ml flask which was filled with sterilized culture medium and shake it thoroughly. You can either leave the filter paper inside the flask or remove it by mean of forceps.

Mix the sediment thoroughly and transfer a spoonful of sediment to another 250 ml flask filled with sterilized culture medium. Rub the leaves in hand with appropriate volume of filtered seawater, filter using a nylon mesh and inoculate 1 ml of the filtrate into the last 250 ml flask which was filled with sterilized culture medium. Incubate the 3 flasks for one week under continuous illumination. Store the test tubes and the remaining of the medium in the refrigerator for the next lab.

What you have in the flasks after one week incubation is a small aquatic ecosystem. It contains : phytoplankton. Some invertebrates. Bacteria and other organisms.