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Applications of adenine phosphoribosyltransferase (APRT) family genes in phytocytomine toxicity aspect

A technology of cytokinin and plant cells, which is applied in the field of application of APRT family genes in plant cytokinin toxicity, can solve the problems of sensitive and invisible cytokinin toxicity, and achieve the effect of reducing toxic and side effects

Inactive Publication Date: 2011-10-19
PEKING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this promotion was not seen in wild-type tobacco, which is sensitive to the toxic effects of cytokinins

Method used

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  • Applications of adenine phosphoribosyltransferase (APRT) family genes in phytocytomine toxicity aspect
  • Applications of adenine phosphoribosyltransferase (APRT) family genes in phytocytomine toxicity aspect
  • Applications of adenine phosphoribosyltransferase (APRT) family genes in phytocytomine toxicity aspect

Examples

Experimental program
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Effect test

Embodiment 1

[0042] Example 1 Application of the Arabidopsis gene APT1 and its homologous genes APT2, APT3, APT4 and / or APT5 in the regulation of plant cytokinin toxicity and cytokinin metabolism

[0043] First, a new screening system for cytokinin mutants was established. Because the previous mutant screening is often screened under low-concentration cytokinin conditions, the mutants of the ethylene synthesis pathway and the ethylene signal transduction pathway are often obtained. For this reason, the present invention has carried out a concentration gradient experiment to find a suitable cytokinin Treatment concentration. Hypocotyl and root elongation of dark-grown wild-type seedlings were strongly inhibited by treatment with high concentrations of zeatin (greater than 100 μM), as did ethylene-insensitive mutants, suggesting a role for cytokinins at this concentration Mainly through a pathway independent of ethylene signal transduction ( figure 1 ). Both the high-concentration cytokin...

Embodiment 2

[0056] Example 2 Application of Arabidopsis gene APT1, its homologous genes APT2, APT3, APT4 and / or APT5 in cultivating adenine and / or cytokinin-regulated transgenic plants

[0057] The full length of APT1, APT2, APT3, APT4 and / or APT5 genes was amplified by PCR method, and constructed on the binary expression vector pDr by molecular cloning method, and then transformed into Arabidopsis thaliana by Agrobacterium infection method to obtain excess Transgenic plants expressing APT1, APT2, APT3, APT4 and / or APT5 genes ( Figure 12 ).

[0058] The results showed that the transgenic plants had different cytokinin resistance. The transgenic plants grown under normal conditions were not significantly different from the wild type, but under the treatment of cytokinin, the phenotypes of the five genes were more differentiated, supporting the functional differentiation of the five genes. APT1 is the most sensitive to cytokinin toxicity, APT3 and APT4 are obviously hypersensitive but we...

Embodiment 3

[0059] Example 3 Knockout of APRT Family Genes to Enhance Plant Cytokinin Resistance or Reduce Toxic Side Effects

[0060] The ethylene-insensitive Arabidopsis mutant ein2-9 tt4 was used as the background for EMS mutagenesis and screening, and it was insensitive to high concentration (300 μM) kinetin. At this concentration, the root development and cotyledon greening process of the wild type were strongly inhibited. A mutant C9 was found to grow normally under this treatment condition ( image 3 ). The corresponding mutation site was found by map-based cloning, and the mutation site of C9 was found to be located in an adenine ribose phosphotransferase APT1 (At1g27450).

[0061] The other three allelic mutants of apt1 (apt1-1, apt1-2, apt1-3) were found to be insensitive to high concentrations of cytokinin, cotyledons could turn green, and roots could elongate ( Figure 4 ). like Image 6 As shown, the dark-grown apt1 mutant was no longer sensitive to zeatin above 30 μM, a...

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Abstract

The invention provides applications of adenine phosphoribosyltransferase (APRT) family genes in aspects of toxicity of cytokinin in plant cells, cytokinin metabolism regulation and control, or resistance selection marking. The APRT family genes comprise Arabidopis thaliana gene APT1 and its homologous genes of ART2, APT3, APT4 and / or APT5. Through an APT1 gene knockout process, toxic and side effects of cytokinin are avoided and plants without APRT gene functions can be utilized as a strain with a toxicity tolerance and for agricultural breeding. The APRT family genes can be utilized for regulation and control of cytokinin metabolism to regulate plant growth and development, such as cell division and differentiation, tissue dedifferentiation and redifferentiation, root cap proportion, leaf senescence, etc. In addition, the APRT family genes can be utilized as resistance selection marking genes. Excessive expressions of APT1 proteins under a treatment condition of high density cytokinin can cause toxicity from metabolism, but APT1 proteins are non-toxic to plants. Therefore, the APRT family genes are in accord with characteristics of selection marking genes and can be utilized for negative selection.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to the application of APRT family genes in the aspect of plant cytokinin toxicity. Background technique [0002] Since the discovery of kinetin in the 1950s, various types of cytokinins have been widely used in agricultural production and tissue culture as an important class of plant growth regulators. Cytokinins commonly used in practice include kinetin and 6-benzyl adenine. As a plant hormone, a small amount of application can have a significant effect. In practice the concentration applied is often much higher than the physiological concentration of cytokinins. On the one hand, it takes into account the solubility and permeability of the drug, and on the other hand, the persistence of the drug effect. However, in the process of application, due to improper application of cytokinin, it often causes toxic side effects of plants. This is a problem that cannot be ig...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29A01H5/00
Inventor 郭红卫张新岩
Owner PEKING UNIV
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