Mild constitutive expression promoter separated from populus tomentosa and application thereof

A technology of constitutive promoter and Populus tomentosa, applied in application, recombinant DNA technology, botanical equipment and methods, etc., can solve the problem of breaking the balance of plant metabolism, unfavorable plant growth, and inability to effectively regulate the target gene in time and space Expression and other issues

Active Publication Date: 2012-06-27
BEIJING FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although the constitutive promoter has brought a lot of convenience to genetic engineering research, it cannot effectively regulate the expression of the target gene in time and space because it drives the constant and continuous expression of the target gene in various tissue parts of the plant, which will cause excessive consumption of cells. Therefore, some problems were exposed during its application (Gittins J R, Pellny T K, Hiles E R, et al.Transgene expression driven by heterologous ribulose-1, 5-bisphosphate carboxylase / oxygenase small-subunit gene promoters in the vegetative tissues of apple(Malus pumila mill.)[J].Planta.2000, 210(2):232-240.), for example: exogenous genes expressed in the whole plant, resulting in a large number of heterologous proteins or metabolites Accumulate in plants, breaking the metabolic balance of plants, which is not conducive to plant growth (Robinson D J.Environmental risk assessment of releases of trangenic plants containing virus-derived inserts[J].Transgenic Res.1996,5:359-362.)
In addition, repeated use of the same constitutive promoter to drive the expression of two or more exogenous genes may cause gene silencing or co-suppression (Kumpatla S P, Chandrasekharan M B, Iyer L M, et al. Genome intruder scanning and modulation systems and transgene silencing [J]. Trends in Plant Science. 1998, 3(3): 97-104.)

Method used

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  • Mild constitutive expression promoter separated from populus tomentosa and application thereof
  • Mild constitutive expression promoter separated from populus tomentosa and application thereof
  • Mild constitutive expression promoter separated from populus tomentosa and application thereof

Examples

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Embodiment 1

[0051] Example 1 Cloning and Analysis of Populus tomentosa Moderate Constitutive Promoter PtMCPpro Sequence

[0052] 1.1 Cloning of Populus tomentosa mild constitutive promoter PtMCPpro sequence

[0053] According to the operating procedure of the Plant Genomic DNA Extraction Kit of Beijing Tiangen Company, the leaves of the aseptic tissue-cultured seedlings of the Populus tomentosa clone LM50 were used as materials to extract genomic DNA, and identified by 1.2% agarose gel electrophoresis. Primers used in PCR amplification include:

[0054] Upstream primer 5'-CTAC GAGCTC TACTAAATAAATATATAA-3',

[0055] Downstream primer 5'-AT GGTACC ATCTATCTGCCCCCTTGTC-3',

[0056] Sac I and Kpn I restriction sites were added to the 5' ends of the upstream and downstream primers, respectively. 20μL PCR reaction system includes: 2μL (100ng) Populus tomentosa genomic DNA, 2μL 10×LA PCR buffer, 1.6μL (2.5mmol / L) dNTP, 0.4μL 10pm / μL upstream primer, 0.4μL 10pm / μL downstream primer, 0.2 μL...

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Abstract

The invention discloses a mild constitutive expression promoter separated from populus tomentosa and application thereof. The mild constitutive expression promoter is a polynucleotide sequence shown by SEQ ID No.1. The invention further discloses a host cell and a recombination plant expression vector comprising the constitutive expression promoter. The constitutive promoter is converted into arabidopsis thaliana after connected with a GUS gene in an operable mode, and a transgene plant is obtained. GUS histochemical staining and GUS activity analysis prove that the separated promoter is a constitutive promoter which leads the GUS gene to carry out mild transcription or expression in various tissues of the plant and each development stage. The invention further discloses application of the constitutive promoter and the recombination plant expression vector comprising the constitutive promoter in the aspects of building transgenic plants, cultivating new plant species, establishing plant bioreactors, analyzing gene functions and the like.

Description

technical field [0001] The invention relates to a promoter isolated from plants, in particular to a mild constitutive expression promoter isolated from Populus tomentosa Carr. and its encoded protein. The present invention also relates to a promoter containing the mild constitutive Recombinant plant expression vectors and host cells expressing promoters. The present invention further relates to their application in the construction of transgenic plants, the cultivation of new plant varieties, the establishment of plant bioreactors, and the analysis of gene functions. It belongs to the category of plant constitutive expression promoters. Separation and its fields of application. Background technique [0002] The promoter is a DNA sequence located in the upstream region of the 5' end of the structural gene, which can activate RNA polymerase to accurately combine with the template DNA and have the specificity of transcription initiation. The promoter determines the direction a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/82C12N1/15C12N1/19C12N1/21C12N5/10C07K14/415A01H4/00A01H5/00
Inventor 安新民陈仲叶梅霞
Owner BEIJING FORESTRY UNIVERSITY
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