A kind of genetic transformation method of Haematococcus pluvialis

A genetic transformation method, the technology of Haematococcus pluvialis, applied in the field of genetic transformation of Haematococcus pluvialls, can solve the problems of low positive transformation rate, low success rate, cumbersome operation, etc., achieve broad market prospects, long cycle, Avoiding the Effects of Cellular Stress Responses

Active Publication Date: 2022-07-15
SHENZHEN UNIV
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Problems solved by technology

[0005] Therefore, the technical problem to be solved by the present invention is to overcome the defects of the genetic transformation method of Haematococcus pluvialis in the prior art, such as cumbersome operation, high cost, low positive transformation rate and low success rate, thereby providing a kind of Haematococcus pluvialis Methods of genetic transformation of algae

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  • A kind of genetic transformation method of Haematococcus pluvialis
  • A kind of genetic transformation method of Haematococcus pluvialis
  • A kind of genetic transformation method of Haematococcus pluvialis

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Embodiment 1

[0061] The present embodiment provides a genetic transformation method of Haematococcus pluvialis, comprising the following steps:

[0062] 1 Linearization of plasmids for transformation

[0063] The plasmid used in this example was pH124. The pH124 plasmid was extracted from Escherichia coli DH5α using Plasmid Mini Kit II (Omega Bio-tek, USA), the plasmid was linearized with restriction endonuclease Not I, and digested with Not I. The system is 30 μL, and specifically includes: 20 μL DNA solution, 2 μL Not I Fast Dicer (New England (NEB), USA), 3 μL Cutsmart Buffer (New England (NEB), USA), 5 μL water. The digestion temperature was 37° C. and the digestion time was 2 h to obtain a linearized pH124 plasmid with the DNA sequence shown in SEQ ID NO: 1, and the linearized plasmid concentration was about 1.0 μg / μL.

[0064] 2 Genetic transformation of Haematococcus pluvialis

[0065] (1) Collect Haematococcus pluvialis cells

[0066] The Haematococcus pluvialis cell selected i...

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Abstract

The invention belongs to the technical field of bioengineering, in particular to a genetic transformation method of Haematococcus pluvialis. The method includes the following steps: using polyethylene glycol solution to mediate the entry of exogenous genes into the protoplast of Haematococcus pluvialis. The present invention first proposes to use polyethylene glycol solution to mediate the transfer of exogenous genes into the protoplast of Haematococcus pluvialis. Since polyethylene glycol can increase the permeability of the cell membrane, the linearized plasmid can be more easily transferred into the pluvialis Haematococcus protoplast, and the method does not cause mechanical damage to cells, avoids cell stress response caused by damage, thereby affecting the transformation of Haematococcus pluvialis. The method solves the problems of complicated operation, high cost, long cycle and low transformation rate in gene gun and Agrobacterium-mediated genetic transformation. The method can be widely used in Haematococcus cell superior character improvement, gene editing and expression regulation, etc., and has huge commercial advantages and broad market prospects.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, in particular to a genetic transformation method of Haematococcus pluvialis. Background technique [0002] Haematococcus pluvialis, as an important economic green algae, is the main source of natural astaxanthin. Astaxanthin is a carotenoid, which can delay aging, promote antibody production and enhance body immunity. It is a natural antioxidant; astaxanthin is also effective in the prevention and treatment of oxidative stress diseases such as diabetes, gout, hypertension and cancer in humans. Therefore, astaxanthin has great market value, and Haematococcus pluvialis has great research significance. [0003] At present, most of the Haematococcus pluvialis used for commercial production of natural astaxanthin are wild-type varieties obtained by screening. The content of astaxanthin in the cells of this wild-type Haematococcus is very limited (below 2%), which can be found in the laboratory...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/82C12R1/89
CPCC12N15/8206
Inventor 胡章立郭春立王潮岗王会
Owner SHENZHEN UNIV
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