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Method for preparing scenedesmusobliquus cell protoplast

A technology for Scenedesmus obliquus and protoplasts is applied in the field of preparing Scenedesmus obliquus protoplasts, which can solve the problem of low efficiency of microalgae protoplasts and achieve the effects of strong activity and strong regeneration ability.

Inactive Publication Date: 2017-10-27
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to solve the problem of low efficiency of preparing microalgae protoplasts in the prior art, and provide an efficient and simple method for preparing Scenedesmus obliquus protoplasts

Method used

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  • Method for preparing scenedesmusobliquus cell protoplast
  • Method for preparing scenedesmusobliquus cell protoplast

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Embodiment 1

[0027] Scenedesmus oblique cells (from the Freshwater Algae Species Bank of the Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan) were cultured in BG11 medium, and the medium composition was: 1.5g / L NaNO 3 , 0.04g / L K 2 HPO 4 , 0.075g / L MgSO 4 .7H 2 O, 0.036g / LCaCl 2 .2H 2 O, 0.006g / L citric acid, 0.006g / L ferric ammonium citrate, 0.001g / L Na 2 EDTA, 0.02g / L Na 2 CO 3 , 2.86mg / L H 3 BO 3 , 1.81mg / L MnCl 2 .4H 2 O, 0.22mg / L ZnSO 4 .7H 2 O, 0.39 mg / L Na 2 MoO 4 .2H 2 O, 0.08mg / L CuSO 4 .5H 2 O, 0.05mg / L Co(NO 3 ) 2 .6H 2 O. The culture conditions are: the temperature is 26±1°C, and the light intensity is 50 μmol m -2 ·s -1 , light cycle 14: 10h condition, culture 10-14 days to the logarithmic growth phase, shake the algae liquid with an ultrasonic oscillator for 30 seconds to break up the cell population formed by the aggregation of Scenedesmus obliques, centrifuge at 2000g, and collect Scenedesmus obliques Cells were resuspended in fresh BG1...

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Abstract

The invention discloses a method for preparing scenedesmusobliquus cell protoplast. The preparation method comprises the following steps: firstly, collecting and culturing a scenedesmusobliquus algae solution during logarithmic growth phase, carrying out oscillating by using an ultrasonic oscillator and then centrifugally collecting scenedesmusobliquus cells; secondly, carrying out resuspending with a BG11 culture medium to obtain scenedesmusobliquus cell suspension; thirdly, adopting mixed digestive enzyme liquid extracted from daphnia magna, adding a small amount of cellulase, pectinase and hypertonic solvent mannitol and membrane stabilizer CaCl2 and then mixing a mixed solution with the scenedesmusobliquus cell suspension, and carrying out enzymatic digestion on the cell wall of the scenedesmusobliquus to obtain the protoplast. In the method disclosed by the invention, the daphnia magna is mixed with digestive enzyme to realize high activity; the cell wall of the scenedesmusobliquus can be subjected to enzymolysis under the mild conditions in a short time. The protoplast prepared by the method has the characteristics that the efficiency can reach 70 percent; in addition, the protoplast has relatively-high activity and high regeneration ability. The invention provides technical support for cell fusion and gene manipulation which aim at scenedesmusobliquus, and also can provide reliable raw materials for studying the endocytosis of microalgae cells.

Description

technical field [0001] The invention belongs to the field of microalgae biotechnology, and relates to a method for preparing Scenedesmus obliquus protoplasts, in particular to a method for obtaining a large amount of active protoplasts by digesting cell walls of algae cells with digestive enzymes of aquatic organisms. Background technique [0002] Microalgae are a class of low-level single-celled organisms that are widely classified in terrestrial and marine ecosystems. Microalgae grow rapidly, because their cells are rich in protein, polysaccharides, polyunsaturated fatty acids, trace elements, pigments and antioxidants and other nutrients, they are widely used in bioenergy, biochemical materials, medicine and health food and aquaculture bait etc. have been widely used, especially microalgae can use solar energy, inorganic salts, CO 2 , Sea water, waste water and non-cultivated land accumulate a large amount of effective biomass, and the development of microalgae resources...

Claims

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Application Information

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IPC IPC(8): C12N1/12C12N13/00C12R1/89
CPCC12N1/12C12N13/00
Inventor 王长海何梅琳颜永全宋虹顾传坤
Owner NANJING AGRICULTURAL UNIVERSITY
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