Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave
A technology of lipase and ultrasound, applied in the field of bioengineering, can solve problems affecting the efficiency of laver cell disruption, unfavorable laver polysaccharide and laver protein extraction, etc.
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Embodiment 1
[0026] Example 1 Preparation of yeast-displayed lipase and yeast-displayed protease
[0027] Synthesize lipase gene (Genbank No.: AF229435), protease gene (Genbank No.: NM_012708.2) and cell wall α-lectin gene of Pichia pastoris GS115 (Genbank No.: M28164) by artificial synthesis. (or protease gene) C-terminus plus linking peptide sequence GSSGGSGGSGGSGGSGS (linker), the nucleotide sequence LIP-linker-α-agglutinin (or PRO-linker-α-agglutinin) is obtained after connection, and EcoR I is added at both ends of the sequence and Not I restriction sites, wherein LIP is the lipase gene (PRO is the protease gene), and α-agglutinin is the cell wall α-lectin gene.
[0028] Using the above-mentioned artificially synthesized sequences as templates, PCR amplification was carried out using the following primer pairs,
[0029] Primer pairs for lipase:
[0030] Upstream primer: 5'-AAGGAAAAAAAGAATTCGTTCCAGTTTCTGG~3';
[0031] Downstream primer: 5'-TTTTCCTTTTGCGGCCGCTAATGAAACG~3'
[0032] P...
Embodiment 2
[0039] Example 2 Yeast Displayed Lipase Collaborative Ultrasonic Disruption of Porphyra Cells
example 1
[0040] Example 1: 1000 grams of water, 20 grams of dried seaweed powder, and 1 gram of yeast-displayed lipase were prepared into a mixed suspension. The stirring speed of the system was 100 rpm, the temperature was 35°C, and the treatment time was 30 minutes. 730W, ultrasonic treatment time of 65min, and treatment temperature of 30°C to obtain cell disruption liquid, centrifuge (4000g 30min) to remove the precipitate, and measure the yield of laver protein and laver polysaccharide in the supernatant.
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