Single cell culture techniques

1. Carefully handle the water used as a culture of algae cultivation as a culture algae cultivation water, must be sterilized by boiling, cooled to room temperature before use, in the cooling process must be covered, no cover container can be covered with clean white paper Bottle mouth. The second and third grades of algae cultivation water can be disinfected chemically, that is, in the pool has been washed into the sand filter or absorbent cotton filtered seawater, and then in the pool of sea water into a certain amount of bleaching powder, The content of available chlorine in the seawater in the pond is 20-30 mg/L, and the mixture is fully mixed and disinfected after 8 to 10 hours, and then the air is inflated. Generally, the air is aerated for about 10 hours, and an equivalent amount of sodium thiosulfate is added. , To neutralize the available chlorine in the water body, and can be used after the determination of residual chlorine in the water body.
The treated seawater can be used up within a short period of time. Try not to exceed one day to prevent it from being contaminated again.
Disinfection of capacity and tools As containers for the cultivation of algae grade flasks, etc., they must be sterilized by heating or boiled disinfection. As glass bottles, cement tanks, etc. for productive expansion cultures, they are usually sterilized with 300-400 mg/L potassium permanganate solution or bleaching powder solution containing 200-300 mg/L available chlorine. Specific methods are: Rinse the container or algae pool with fresh water or seawater first, and then soak the container or tool in the prepared potassium permanganate solution (or bleach solution) for 5 to 10 minutes. Remove and rinse with disinfected seawater. Clean and ready to use. To disinfect the algae pool, you can use the prepared potassium permanganate solution (or bleach solution) to be showered from the top of the pool wall and splash the bottom of the pool. After about 10 minutes, rinse with sterile seawater to use.
Third, increase the proportion of inoculation in all levels of culture The general algae species-level culture access to the ratio of the algae species and the new culture fluid is 1:0.5 to 1, intermediate or large-area culture, their ratio is 1:3 to 6 Therefore, the number of reproducible monocytic algae in the algal fluid is large, and the algae in the culture fluid have a rapid growth rate, which shortens the cultivation time and relatively reduces the pollution. On the other hand, antagonism between organisms is also used to increase the concentration of algae in the culture medium to inhibit the proliferation of other algae and predator organisms and to reduce the possibility of contaminating the algal fluid.
IV. Inoculation time The inoculation time is generally selected between 8:00 and 10:00 in the morning. It is not appropriate to vaccinate at night because many algae sink to the bottom at night. During the day, algae carry out photosynthesis, and the light tends to rise, making it easier to observe the movement. Ability, but also play a role in selection.
Fifth, daily management should pay attention to stirring and stirring can promote the growth and reproduction of algae, its role is mainly:
First, increase the contact surface of water and air by stirring to supplement the consumption of carbon dioxide due to the photosynthesis of algae;
Second, help the precipitated algae cells to float and gain light;
Third, prevent pellicles from forming on water surfaces. Stirring should be done regularly and change direction often to avoid swirling water flow. Do not stir too much at the same time to prevent splashing of algae.
Illumination of light is suitable or not, and it has a great influence on the growth of cultivated algae. At present, the production units are all cultivated using natural light, but they are volatile. Therefore, in the cultivation and management, the intensity of light must be adjusted according to the weather, and the light intensity should be maximized. May be suitable for cultivation of algae requirements. The sunny algae room is shaded with white Dacron cloth as a curtain to prevent direct light from shining; the light is worst in rainy days. The culture bottle should be placed as close as possible to the window. Conditional light sources can be used.
Temperature regulation Each kind of algae has its adaptable temperature range and optimum temperature range, but at present, most of the culture of monocytic algae do not have temperature control conditions and can only follow the change of natural temperature, and can adapt to or adapt to different seasons. Basically adapt to the local temperature changes in the cultivation of species.
In general, the high temperature is harmful to the growth of algae. In indoor cultivation in the summer, the doors and windows must be opened for ventilation and cooling.
VI. Predator Control In the cultivation of monocytic algae, the pollution and hazards of predator organisms are the main reasons for the failure of algae culture. The enemy should be "prevented, prevention and control combined," to minimize its harm. Prevention measures should be applied throughout the algae cultivation process.
First, it strives to ensure the purity of algae species and nutrient salts and control source pollution.
Secondly, seawater must be strictly filtered and disinfected. Instruments and algae culture ponds must also be strictly disinfected. At the same time, the proportion of inoculation should be increased to suppress the proliferation of other algae and other predator organisms, and reduce the possibility of contaminated algae.
In addition, in the daily management, keep the algae room clean and sanitary to prevent dust and insects from being contaminated.
Elimination, suppression or killing of predatory biological methods:
One is the use of filtering methods to remove large enemy predators. The unicellular algae are very small and can be removed by the filtering method on contaminated large enemy organisms (such as rotifers). Usually rotifers can be screened with sieve filters smaller than 60 μm, and once filtered, adult rotifers can be removed. However, rotifer eggs and developing young individuals cannot be completely removed. Therefore, they must be continuously filtered for 3 days, once a day. The eggs and young individuals that survived the first filtration develop into adults and are removed during the second or third filtration.
The second is the use of drugs to suppress or kill the enemy. The drugs used to suppress or kill the enemy's harmful organisms include chemical drugs and Chinese herbal medicines. However, due to the fact that predator organisms are more resistant to drugs than algae, the effect is not ideal. At present, there are not many effective drug methods. Although some preliminary effects can be achieved by killing predator organisms, further tests are needed.
The third is to change environmental conditions to kill predators. The use of this method must understand the range of adaptation of the cultivated algae and predator organisms to the ecological factors, and then according to the specific circumstances, change an environmental factor, to achieve the purpose of killing the enemy and preserving algae.

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