Sterilizing method of spherical phaeocystis culture solution

A technology of Phaeocystis globosa and culture solution, which is applied in the field of microalgae culture solution, can solve the problem of not being able to detect whether there are bacteria in the algae solution, and achieve the effect of simple operation and high efficiency

Inactive Publication Date: 2012-03-28
XIAMEN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In previous experiments, we found that eubacterial 16S rDNA conservative primers can amplify the mitochondrial DNA fragments of Phaeocystis globosa, so this method cannot be used to detect whether there are bacteria in the algae liquid

Method used

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  • Sterilizing method of spherical phaeocystis culture solution
  • Sterilizing method of spherical phaeocystis culture solution
  • Sterilizing method of spherical phaeocystis culture solution

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1: Testing the tolerance of Phaeocystis globosa to SDS

[0029] (1) Dispense the Phaeocystis globosa algae solution grown to the logarithmic phase in a 24-well cell plate, 1.8 mL per well;

[0030] (2) Dilute SDS (10%, m / v) with f / 2 medium to a series of concentration gradients, take 200 μL and add it to 1.8 mL algae liquid, so that the final concentrations of SDS in the algae liquid are: 0.01%, 0.005% , 0.0025%, 0.001%, 0.0005% (m / v), 200 μL f / 2 medium was added in 1.8mL algae liquid as a control, and each treatment concentration was set to repeat three times;

[0031](3) Cultivate under light at 20°C for 5 days;

[0032] (4) After 5 days, each well was mixed, and 200 μL of algae liquid was taken from it in a 96-well microplate, and 200 μL of algae medium was used as a blank control, and their fluorescence values ​​were detected with a microplate reader (excitation wavelength 440nm, emission wavelength 680nm) ;

[0033] (5) Use the following formula to calcu...

Embodiment 2

[0040] Example 2: Testing the tolerance of Phaeocystis globosa to antibiotics

[0041] (1) Select 8 kinds of antibiotics and test their influence on the growth of Phaeocystis globosa respectively. Namycin 1mg / mL, streptomycin 10mg / mL, cephalosporin 5mg / mL, ampicillin 10mg / mL, rifamycin 1mg / mL;

[0042] (2) Dispense the Phaeocystis globosa algae solution grown to the logarithmic phase in a 24-well cell plate, 1.8 mL per well;

[0043] (3) Dilute the antibiotic mother solution by 10 times and 100 times with f / 2 medium, take 200 μL of the mother solution and the dilution solution and add it to 1.8 mL of the algae liquid, so that the final concentrations of the antibiotics in the algae liquid are 10%, 1%, and 10% of the mother liquid respectively. 0.1%, 200μL f / 2 medium was added to 1.8mL algae liquid as a control, and 3 replicates were set for each treatment concentration;

[0044] (4) Cultivate under light at 20°C for 5 days;

[0045] (5) After 5 days, each well was mixed eve...

Embodiment 3

[0053] Embodiment 3: the sterilization of Phaeocystis globosa algae liquid

[0054] (1) Take 20mL of Phaeocystis globosa liquid grown to the logarithmic phase, centrifuge at 2000rpm for 5min, and remove the supernatant;

[0055] (2) Resuspend the algae cells with 20mL f / 2 medium, centrifuge at 2000rpm for 5min, remove the supernatant, repeat this step twice, and then resuspend the algae cells with 20mL f / 2 medium;

[0056] (3) Add SDS (final concentration is 0.0025%) and 8 kinds of antibiotics simultaneously (final concentration is: clindamycin 5mg / L, azithromycin 50mg / L, gentamicin 100mg / L, kanamycin 100mg / L , streptomycin 1000mg / L, cephalosporin 500mg / L, ampicillin 1000mg / L, rifamycin 1mg / L);

[0057] (4) Cultivate under light at 20°C for 5 days. During this period, take 2 mL of the treated algae solution every day, centrifuge at 2000 rpm for 5 minutes, remove the supernatant, resuspend the algal cells with 2 mL of f / 2 medium, centrifuge at 2000 rpm for 5 minutes, remove th...

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Abstract

The invention provides a sterilizing method of a spherical phaeocystis culture solution, and relates to a microalgae culture solution. The method comprises the following steps: centrifuging algae solution which grows to an exponential phase, and removing supernatant; carrying out gravity suspension on algae cells with an f / 2 culture medium, centrifuging, and removing supernatant, repeating twice, and carrying out gravity suspension on the algae cells with the f / 2 culture medium; adding SDS (sodium dodecyl sulfate) and antibiotics, wherein the antibiotics are clindamycin, azithromycin, gentamicin, kanamycin, streptomycin, cefotaxime, ampicillin and rifamycin; culturing under illumination, centrifuging the algae solution and removing supernatant during culturing, carrying out gravity suspension on the algae cells with the f / 2 culture medium, centrifuging and removing supernatant, removing residual SDS and the antibiotics, transferring into the f / 2 culture medium, and culturing under illumination; and selecting survival transferred algae solution which is treated with SDS and the antibiotics, and detecting whether bacteria exist or not after the transferred algae solution grows to the exponential phase. The method is convenient to operate, complex operations, such as bacterium separation and test on sensitivity to antibiotics and the like, are avoided, and long-term treatment of high-concentration antibiotics has good sterilization effect.

Description

technical field [0001] The invention relates to a microalgae culture solution, in particular to a sterilization method for the Phaeocystis globosa culture solution. Background technique [0002] In the research field of microbial control of red tide, the existence of bacteria in the algae liquid brings great difficulties to the purification of unculturable algicidal microorganisms, as well as the research on the algicidal mechanism of algicidal microorganisms and algicidal substances, so it is hoped that through The algae liquid is sterilized to eliminate the interference of bacteria in the algae environment. [0003] The currently reported algae liquid sterilization methods mainly include: ①Pick a single algal cell under a microscope, wash it repeatedly with a sterile algae medium, and then inoculate it in the algae medium. This method is cumbersome to operate and requires a certain amount of algae separation At the same time, it is difficult to cultivate a single algae ce...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/12C12R1/89
Inventor 郑天凌黄丽萍苏建强郑小伟周艳艳
Owner XIAMEN UNIV
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