Dunaliella salina CPD photolyase, photolyase liposome prepared from the same and preparation method for the photolyase liposome

A technology of Dunaliella salina and photolytic enzyme lipid, applied in the field of biology, can solve problems such as production reduction, and achieve a good effect of repairing ultraviolet damage

Inactive Publication Date: 2008-03-19
SICHUAN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The main harm of UV-B to plants is to inhibit their p...

Method used

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  • Dunaliella salina CPD photolyase, photolyase liposome prepared from the same and preparation method for the photolyase liposome

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Experimental program
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Effect test

Embodiment 1

[0021] Cloning of the CPD photolyase gene from Dunaliella salina.

[0022] 1. Collection of Dunaliella salina

[0023] Dunaliella salina was purchased from the algae bank of Wuhan Institute of Hydrobiology, Chinese Academy of Sciences.

[0024] 2. Poly A+RNA isolation (Poly A+RNA isolation)

[0025] Total RNA of Dunaliella salina was extracted with Trizol reagent (Invitrogen, USA). The quality of total RNA was identified by formaldehyde denaturing gel electrophoresis.

[0026] 3. Synthesis of cDNA by RT-PCR

[0027] Using a reverse transcription kit (TaKaRa RNA PCR Kit (AMV) ver.2.1), follow the instructions to reverse transcribe RNA into cDNA.

[0028] 3. Full-length cloning of Dunaliella salina CPD photolyase gene (Cloning of Full-length cDNA)

[0029] According to the amino acid conserved sequence of CPD photolyase gene of known species (such as Chlamydomonas, Arabidopsis, cucumber, spinach, rice, etc.), degenerate primers were designed, and the principle of homologous...

Embodiment 2

[0042] Sequence Analysis of CPD Photolyase Protein from Dunaliella salina

[0043] The full-length sequence of Dunaliella salina CPD photolyase and its coding sequence were subjected to nucleotide and amino acid BLAST on NCBI, and it was found that there was no obvious sequence homology in the nucleic acid database. At the amino acid level, it has 52% identity with Chlamydomonas reinhardtii CPD photolyase, 48% identity with cucumber (Cucumissativus) CPD photolyase, and rice (Oryza sativa) CPD photolyase. 47% identity, and 49% identity with Arabidopsis thaliana CPD photolyase.

Embodiment 3

[0045] Construction of Expression Vector of CPD Photolyase Gene of Dunaliella salina

[0046] According to the full-length coding sequence of Dunaliella salina CPD photolyase gene, primers were designed to amplify the complete coding reading frame, and restriction enzymes BamHI and SalI restriction sites were introduced into the forward and reverse primers respectively. After PCR amplification, the Dunaliella salina CPD photolyase cDNA was connected to the vector pMD18-T, then digested with BamHI and XhoI, and further connected to the prokaryotic expression vector pGEX-4T-1. Then the constructed expression vector Transformed into Escherichia coli ultraviolet damage repair-deficient strain SY2 competent cells, picked a single clone, carried out enzyme digestion and PCR verification, the results proved that the Dunaliella salina CPD photolyase gene expression vector was successfully constructed.

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Abstract

The present invention provides dunaliella salina CPD photoreactivating enzyme which can rehabilitate the damage of ultraviolet radiation, a nucleotide sequence which codes the dunaliella salina CPD photoreactivating enzyme and the photoreactivating enzyme plastid which is prepared by the dunaliella salina CPD photoreactivating enzyme and can rehabilitate the damage of ultraviolet radiation,; wherein, the prepared photoreactivating enzyme plastid can be used as the raw material which is used to produce the medicine which rehabilitates the damage of ultraviolet radiation, and sun screening agent. The present invention also provides preparation methods of the dunaliella CPD photoreactivating enzyme and the photoreactivating enzyme plastid which can rehabilitate the damage of ultraviolet radiation.

Description

1. Technical field [0001] The present invention belongs to the field of biology. Specifically, the present invention relates to the photolyase gene and protein related to ultraviolet damage repair, the application of enzyme protein in the preparation of ultraviolet damage repair liposomes, and the application of genes, proteins and ultraviolet damage repair liposomes. method of preparation. 2. Technical background [0002] With the rapid development of world industry, the emission of a large amount of greenhouse gases leads to global warming, especially the extensive use of chemical substances such as chlorofluorocarbons (chlorofluorocarbons, chlorofluorocarbons, CFCs), which makes the ozone layer in the atmosphere thinner and thinner, resulting in Terrestrial solar radiation (especially UV-B, 280-320nm) is increasing day by day. The increase of solar UV-B radiation intensity near the surface will have a profound impact on various organisms living in the earth's biosphere a...

Claims

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Application Information

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IPC IPC(8): C12N15/52C12N9/00
Inventor 曹毅乔代蓉白林含徐辉程龙
Owner SICHUAN UNIV
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