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Chinese cabbage 9-cis-carotene epoxide dioxygen zymoprotein coding sequence

A technology of dioxygenase protein and carotene, applied in the field of genetic engineering, can solve problems that have not been discovered and pointed out

Inactive Publication Date: 2004-09-15
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In the analysis of existing literature, although "The Plant Journal, 2001, 27: 325-333" has fully affirmed the function of NCED protein, pointing out that the gene can increase the expression of ABA under drought stress, but did not point out The expression of this gene is related to adversities such as salt and ultraviolet rays, and so far, no reports have been found that are closely related to the subject of the present invention.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0185] Cloning of 9-cis-epoxycarotenoid peroxidase gene from Chinese cabbage

[0186] 1. Tissue separation (isolation)

[0187] Chinese cabbage (the variety is "Heat-resistant No. 3") was purchased from the market, and the Chinese cabbage was germinated at 28°C for 24 hours, and then sowed in the greenhouse. When the leaves of the Chinese cabbage were 4-5 pieces, it was ready to extract DNA or RNA.

[0188] 2. RNA isolation (RNA isolation)

[0189] Take part of the tissue, grind it with a mortar, add it to a 1.5mL EP tube filled with lysate, shake it fully, and then transfer it into a glass homogenizer. After homogenization, transfer to 1.5mL EP tube, and extract total RNA (TRIzol Reagents, GIBCOBRL, USA). The quality of total RNA was identified by formaldehyde denaturing gel electrophoresis, and then the RNA content was determined on a spectrophotometer.

[0190] 3. Cloning of Full-length cDNA

[0191] According to the amino acid conserved sequence of Arabidopsis thalian...

Embodiment 2

[0203] Sequence information and homology analysis of 9-cis-epoxycarotenoid peroxidase-related genes in Chinese cabbage

[0204] The length of the full-length cDNA of the new Chinese cabbage 9-cis-epoxycarotenoid peroxidase of the present invention is 1151bp, and the detailed sequence is shown in SEQ ID NO.3, wherein the open reading frame is located at 89-1774 nucleotides (699 nucleotides). The amino acid sequence of Chinese cabbage 9-cis-epoxycarotenoid peroxidase was deduced according to the full-length cDNA, with a total of 233 amino acid residues, a molecular weight of 167.63 kDa, and an isoelectric point (pI) of 4.89. See SEQ ID NO.3 for the detailed sequence.

[0205] The full-length cDNA sequence related to Chinese cabbage 9-cis-epoxycarotenoid peroxidase and its encoded protein were published in Non-redundant GenBank+EMBL+DDBJ+PDB and Non-redundant GenBank CDS translations+PDB+SwissProt using BLAST program Nucleotide and protein homology searches in the +Superdate+PI...

Embodiment 3

[0207] Eukaryotic expression of Chinese cabbage 9-cis-epoxycarotenoid peroxidase-related protein or polypeptide in Arabidopsis and identification of stress resistance of transgenic plants

[0208] Containing the construction of the expression vector of target gene (Chinese cabbage 9-cis-epoxy carotenoid dioxygenase gene), according to the full-length sequence (SEQ ID NO.3), design primers to amplify the complete coding reading frame, and introduce restriction endonuclease sites on the upstream and downstream primers respectively (this can be determined by the vector selected), so as to construct the expression vector. Using the amplified product obtained in Example 1 as a template, after PCR amplification, the Chinese cabbage 9-cis-epoxy carotenoid peroxidase gene cDNA was cloned into an intermediate vector (such as pBluescript), and further cloned into a double Meta-expression vectors (such as pBI121 and improved pCAMBIA2300), under the premise of ensuring the correct reading...

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PUM

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Abstract

The invention is a coding series of celery cabbage 9- series- epoxy caritinoid oxydol enzyme protein, which belongs to gene project field. It includes: codes the nucleotide series of polypeptide which has celery cabbage 9- series- epoxy caritinoid oxydol enzyme protein activity, and at least 70% of the series has consanguinity with SEQ ID NO.3 nucleotide from 89 to 1774; or the series can cross with the 89-1774 bit nucleotide series of SEQ ID NO.3. the product in the invention has key effect to the ABA synthsis, it can upgrades the inversion resisting property of plant effectively.

Description

technical field [0001] The invention relates to a protein coding sequence, in particular to a Chinese cabbage 9-cis-epoxy carotenoid dioxygenase protein coding sequence, which belongs to the field of genetic engineering. Background technique [0002] Many experiments have shown that abscisic acid (abscisic acid for short) is a kind of stress hormone, which plays an important role when plants resist external environmental stress, such as heat, salt, drought, cold and other environmental conditions and herbicides, etc. can affect ABA in plants. levels in the body. In some developmental processes of plants, such as seed germination, embryonic development and fruit ripening, the ABA content will change to a certain extent. ABA plays an important role in plant growth and development. In ABA biosynthesis, in addition to the induction of external environmental conditions, some factors in plants, such as enzymes and coenzyme factors, also play a more important regulatory role. In...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H1/00A01H5/00C12N9/02C12N15/53C12N15/79
Inventor 李柱刚唐克轩赵凌侠钱虹妹孙小芬张利达
Owner SHANGHAI JIAO TONG UNIV
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