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A multi-stress-inducible promoter pg5 derived from Eucalyptus grandis and its application

A technology of promoter and Eucalyptus grandis, applied in the field of genetic engineering, can solve problems such as inhibiting plant growth and development, and achieve the effect of saving money and improving the ability to resist various adversities

Inactive Publication Date: 2015-12-02
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these promoters can enable the high-efficiency expression of foreign genes in plants, although the stress resistance of transgenic plants has been improved, they often inhibit the growth and development of plants.

Method used

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  • A multi-stress-inducible promoter pg5 derived from Eucalyptus grandis and its application
  • A multi-stress-inducible promoter pg5 derived from Eucalyptus grandis and its application
  • A multi-stress-inducible promoter pg5 derived from Eucalyptus grandis and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] promoter discovery

[0033] Genomic DNA of Eucalyptus grandis (variety EG6) was extracted, and the extraction of Eucalyptus genomic DNA referred to conventional methods in the art. Design specific primers (P1: 5'-TCAT TCTAGA GCGTAATTGCTGTCGCACCC-3', such as SEQ ID NO: 2, the underline is the XbaI restriction site) and (P2: 5'-GCTC AAGCTT CGAGAAGGAATCGCTTCTGA-3', such as SEQ ID NO: 3, the underline is the SacI restriction site) to amplify the genomic DNA, recover and clone the fragment, and perform DNA sequencing to obtain a 720bp DNA fragment, the nucleotide of the 720bp DNA fragment The sequence is shown in SEQ ID NO:1. The amplified sequence shown in SEQ ID NO: 1 was analyzed for the regulatory region, the start codon ATG of the gene was identified, and the part of the structural gene was removed. Obtain the regulatory sequence at the 5' end of the gene, the regulatory elements of the promoter such as figure 2As shown, the TATA box is located in the region?150-...

Embodiment 2

[0035] 1. Low temperature and drought-induced activity verification of the promoter pG5

[0036] Construction of recombinant expression vector

[0037] 1. Digest pGEM-T: pG5 prepared in Example 1 with restriction endonucleases XbaI and SacI, and recover the digested product (pG5).

[0038] 2. Digest the binary vector pBI121 with restriction endonucleases XbaI and SacI to obtain the pBI121 vector backbone with the CaMV35S promoter removed.

[0039] 3. Ligate the enzyme-digested product of step 1 with the vector backbone of step 2 to obtain a recombinant plasmid, and perform sequencing verification. The sequencing results show that the target plasmid pG5-GUS (replacing the XbaI of pBI121 with pG5 of the sequence shown in SEQ ID NO: 1) and the small fragment between the SacI recognition site).

[0040] 2. Transformation of tobacco by transient expression

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Abstract

The invention belongs to the technical field of genetic engineering, and specifically discloses a multi-stress-inducible promoter pG5 derived from Eucalyptus grandis and its application. The promoter is specifically the promoter of the multi-stress inducible gene G6PDH of the Eucalyptus grandis variety EG6. The nucleotide sequence of the promoter such as SEQ? ID? NO: 1 shown. The regulatory elements of the promoter of the present invention include elements related to low temperature stress response, drought stress response, light response, hormone response and the like. The promoter can be induced and expressed by adversities such as low temperature and drought, can be used as an inducible expression vector of plant origin, and has wide applicability and good application prospects in the field of bioengineering.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a multi-stress-inducible promoter pG5 derived from Eucalyptus macrophylla and its application. Background technique [0002] The pentose phosphate pathway is an important pathway for sugar metabolism in plants. Its main physiological function is to provide reducing power NADPH for biosynthesis and five-carbon sugar for nucleic acid synthesis, and some intermediate products can participate in the synthesis of amino acids and fatty acids, etc. Glucose-6-phosphate dehydrogenase is its key regulatory rate-limiting enzyme (Lin Yuanzhen. 2006. Gene cloning, structural analysis and functional identification of sweet poplar glucose-6-phosphate dehydrogenase. Doctoral dissertation, Beijing Forestry University , Supervisor: Zhang Zhiyi, pp.44-66), involved in the plant response caused by a variety of environmental stresses, such as the interaction with some metal ions (such as ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/63C12N1/15C12N1/19C12N1/21C12N5/10A01H5/00
Inventor 林元震莫晓勇杨小红杨会肖周玮陈晓阳
Owner SOUTH CHINA AGRI UNIV