1. Marine Biotechnology Lab
Algal Culture Media

2. content Algal Culture Media
Collection-and-handling-samples-for-Algal-isolation
Protocols for Isolation and Purification of a single Algal Species
Algal Culture Methods
Monitoring algal populations
Production and Application of Rotifers in Aquaculture
Production of the brine shrimp Artemia
Determination of active chlorine content
Preparation of a disinfecting hypochlorite solution
Bioluminescence

3. Introduction Chlorophyta (Green). Rhodophyta (Red).
Algae are a very diverse group of photosynthetic eukaryotic organisms that fall into three major groups:
Chlorophyta (Green).
Rhodophyta (Red).
Phaeophyta (Brown).

4. In their natural habitats algae obtain all the nutrients, minerals and vitamins they require from the water in which they live.
To grow them in the lab however, you must provide them with all of these essential resources i.e. you will need to make up some growth media.

5. All the media have several components in common:
Algae media
refers to the solution or culture in which algae grow.
All the media have several components in common:
sources of nitrogen (in the from of nitrate, nitrite and ammonia),
sources of phosphorus
sources of vitamins
sources of trace metals.
However the specific types of these nutrients, their concentrations and ratios vary between the media.

6. There are many recipes for such algal growth media :
AF6 Medium
Black Sea Medium
f/2 Medium
Freshwater Test Medium
K Medium
Modified K Medium

7. F/2 medium
Is the most common and widely used general enriched seawater medium designed for growing marine algae used in the phycological and aquaculture studies.

8. Stock solutions and salts
In any recipes you will see two types of stocks
“working stocks” .
“primary stocks”.
Working stocks
Are those whose aliquots contribute directly to making the final media.
Primary stocks
Are normally made where several single substance solutions are then combined to form the working stock.

9. Stock solutions are made up by accurately weighing the prescribed amount of nutrient and dissolving in a specified volume of distilled water, if possible in a volumetric flask.
Some nutrients will readily dissolve, others need heat and stirring to fully dissolve.
In contrast vitamin stocks are heat sensitive and should not be subjected to heat treatment and should also be kept in the dark.
Failure to fully dissolve the primary stocks of some nutrients such as EDTA can lead to gross precipitation when these stocks are combined to make the media.

10. Nutrients come with different salts and hydration
Nutrients come with different salts and hydration. For example, while copper and zinc may be two desired active constituents they are readily obtained from suppliers with either SO4 or Cl2 salts (ie CuSO4 or CuCl2 and ZnSO4 or ZnCl2).
Some nutrients also come with different hydrations, ie the .nH2O suffix.
Substituting one form for another may have no effect on the growth of some microalgae species, but it can lead to poor growth in others and also lead to unwanted and time consuming precipitation problems as the overall ratio of salts in the medium has changed.
Therefore deviating from the prescribed recipes is to be avoided and ordering the correct form is recommended.

11. Seawater source and treatment
The marine microalgae species should be grown using:
Unpolluted oceanic seawater
Artificial seawater.
Artificial seawater media is composed of marine salts and nutrients added to pure freshwater.
Artificial seawater is only necessary where a clean natural seawater source is unavailable or in particular research studies where the exact composition needs to be controlled.

12. Off-shore sites have very low concentrations of metal and organic pollutants therefore it suitable as the base medium for a wide range of marine microalgae species.
The seawater should be collected in clean black polyethylene containers and then stored until needed (preferred at 4 0C). Then it is treated using a filtration.

13. F/2 Medium Stock Solutions:

14. Mineral salts working solution
Add Nitrate directly to filtered seawater and autoclave.
Use: 1 ml per litre of seawater medium.

15. To 950 mL distilled H2O add: Trace Metal Solution
Solution will initially be cloudy. Add 1N NaOH to adjust pH to about 4.5; solution
should clear unless too much NaOH has been added. Bring final volume to one liter.
Use: 1 ml per litre of sterilized seawater medium.

16. Vitamin Solution
First, prepare primary stock solutions for vitamin B12 and biotin (Vitamin H) according to the proportions indicated below.
To prepare final vitamin solution, begin with 950 mL of dH2O, dissolve the thiamine, add the amounts of the primary stocks as indicated in the quantity column below, and bring final volume to 1 liter with dH2O.
Warning: do not autoclave any vitamin solution but Filter sterilize

17. Store primary stock solutions in freezer
Store primary stock solutions in freezer. Store completed vitamin solution in the refrigerator or freeze small aliquots.
Use: 0.5 ml per litre of sterilized seawater medium.