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High-density culture method of prorocentrum

A technology of high-density culture and Prototheca, applied in microorganism-based methods, biochemical equipment and methods, single-cell algae, etc., can solve the problems of high price, small capacity of light incubator, and high energy consumption of temperature control system, To achieve the effect of simple operation, efficient acquisition, and small culture volume

Active Publication Date: 2018-07-06
EAST CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The light incubator has small capacity and high price, while the walk-in incubator requires high ener

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] In a sterile operating room, suck 19.0L of sterilized f2 medium with a salinity of 28 into a 20L jacketed glass reactor with a sterile silicone tube, and inoculate a concentration of 5.0×10 at a volume ratio of 5% 6 cells / L Prorocentrum PM03, that is, add 1.0L of algae seed solution. Transfer the reaction kettle to a conventional laboratory, use an air pump to connect to a sterile 0.2μm filter and a silica gel tube to vent 3L / min, the speed of the reaction kettle is 80rpm / min, the light is 5000lux, the light / dark cycle is 14h / 10h, and the jacket circulating water temperature Control 22-25°C and cultivate for 15 days. Then in the sterile operating room, most of the cultured algae liquid is discharged from the reactor outlet, and a certain proportion of the volume is reserved for inoculation in the next culture cycle. Using the Sedgewick-Rafter plate to count the cells under the microscope, the cell density of the cultured algae solution was 8.97×10 7 cells / L.

Embodiment 2

[0026] Continuation of Example 1. In a sterile operating room, the cell density retained in a 20L jacketed glass reactor is 8.97×10 7 1.5L of Prorocentrum PM03 seed solution in cells / L, inhaled with a sterilized silicone tube and added 18.5L of sterilized f2 medium with a salinity of 25. Transfer the reaction kettle to a conventional laboratory, use an air pump to connect to a sterile 0.2μm filter and a silica gel tube to vent 3L / min, the speed of the reaction kettle is 80rpm / min, the light is 4000lux, the light / dark cycle is 14h / 10h, and the jacket circulating water temperature Control 22-25°C and cultivate for 15 days. Then in the sterile operating room, most of the cultured algae liquid is discharged from the reactor outlet, and a certain proportion of the volume is reserved for inoculation in the next culture cycle. Using the Sedgewick-Rafter plate to count the cells under the microscope, the cell density of the cultured algae solution was 1.38×10 8 cells / L.

Embodiment 3

[0028] Continuation of Example 2, in a sterile operating room, the cell density retained in a 20L jacketed glass reactor is 1.38×10 8 Cells / L of Prorocentrum PM03 seed solution was 2.0L, and 18.0L of sterilized f2 medium with a salinity of 25 was added to the sterilized silicone tube. Transfer the reaction kettle to a conventional laboratory, use an air pump to connect to a sterile 0.2μm filter and a silicone tube to vent 3L / min, the speed of the reaction kettle is 80rpm / min, the light is 3000lux, the light / dark cycle is 14h / 10h, and the jacket circulating water temperature Control 22-25°C and cultivate for 15 days. Then in a sterile operating room, discharge most of the cultured algae solution 18L from the reactor outlet, and use the Sedgewick-Rafter plate to count the cells under the microscope to find that the cell density of the cultured algae solution is 2.05×10 8 cells / L; The remaining 2.0L of algae seed solution is used for inoculation in the next culture cycle, add 18L o...

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PUM

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Abstract

The invention discloses a high-density culture method of prorocentrum. The high-density culture method comprises the following steps: (a) in an aseptic manipulation room, inoculating prorocentrum seedliquid into a jacketed glass reactor with the volume of 10 to 50 L filled with a sterile f2 culture medium, wherein the salinity of the f2 culture medium is 20 to 28; (b) transferring to a routine test laboratory, introducing sterilized air, controlling the light intensity at 3000 to 5000lux and light/dark cycle at 14h/10h, controlling circulating water temperature of a jacket at 22 to 25 DEG C and culturing for 10 to 20 days; (c) in the aseptic manipulation room, discharging most cultured algae liquid from a discharge opening of a reaction still, adding a fresh sterilized f2 culture medium into the reaction still, transferring to the routine test laboratory and continuously carrying out next culture cycle by using identical conditions in step b. The high-density culture method disclosedby the invention has the advantages that the whole culture process is simple in operation, culture volume is small, cell density is extremely-high and total extracts of the prorocentrum can be obtained more efficiently; in addition, compared with large volume glass bottle or glass jar culture, the high-density culture method has the advantages that the prorocentrum does not need to be cultured inan aseptic and constant temperature culture room, so that energy consumption is low.

Description

Technical field [0001] The invention belongs to the field of culturing marine miniature unicellular algae, and particularly relates to a method for high-density culturing of Prorocentrum. Background technique [0002] Marine organisms are rich in secondary metabolites with novel structures and significant biological activities, making them the most promising source of new drugs. In addition to marine invertebrates such as sponges, corals, and ascidians, and marine microorganisms such as actinomycetes and fungi, Marine microalgae resources are also one of the hotspots in marine natural medicinal chemistry research; the polyketone, polyether, and alkaloid components produced by their metabolism are classified as neurotoxic shellfish due to their specific toxicological characteristics to mammals , Paralytic shellfish poisoning, diarrheal shellfish poisoning, cyclic imine toxins and other 8 major marine biological toxins, their monitoring is the core content of aquatic product qualit...

Claims

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Application Information

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IPC IPC(8): C12N1/12C12R1/89
CPCC12N1/12
Inventor 樊成奇田晓清乔玉宝唐莹莹陆亚男马丽艳
Owner EAST CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI