A method for regulating cell pathways by using plant hormone GA and small molecular substance pac

A technology of small molecular substances and plant hormones, applied in the field of biological method invention, can solve the problems of immune system damage and incompatibility

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Problems solved by technology

However, both this method and optogenetics have certain limitations
Rapamycin is an immunosuppressor, which has certain damage to the immune system; the optogenetics system is not suitable for the regulation of intracellular light signaling pathways

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  • A method for regulating cell pathways by using plant hormone GA and small molecular substance pac
  • A method for regulating cell pathways by using plant hormone GA and small molecular substance pac
  • A method for regulating cell pathways by using plant hormone GA and small molecular substance pac

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Embodiment 1

[0037]A method for regulating cell pathways by utilizing the plant hormone GA and the small molecular substance PAC, comprising the following methods:

[0038]

[0039] (1) Carrier design and construction:

[0040] First, using the primers described above, using Arabidopsis thaliana cDNA and HEK293T cell cDNA as templates, PCR technology is used to amplify each gene fragment, and then using the corresponding primers, using two fragments as templates, carry out overlap PCR; The Arabidopsis DELLA protein GAI and the GA receptor GID1 were fused with the C-terminus of the key protein LRP6 in the Wnt signaling pathway, and the PCR products such as GAI, GID1, GAI-LRP6C, GID1-LRP6C were recovered by agarose gel, and the internal The linearized pCI 4xmyc, pCI egfp and pCI mcherry vectors were digested with XbaI and XmaI enzymes, ligated by Infusion, transformed into DH5a, verified by colony pcr, picked and sent for sequencing, and ensured that the bacteria were correctly sequenced;...

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Abstract

The invention provides a method for regulating cell pathway by using plant hormone GA and small molecular substance PAC, and realizes the regulation switch of signal pathway in human cells. The GA receptor proteins GID1 and GAI in Arabidopsis thaliana were constructed on a human cell expression vector, and then transferred into HEK293T cells together. GA was added to induce the interaction between the two proteins. After adding the inhibitor PAC, the degree of interaction would be weakened. In this way, the key proteins in the human cell signaling pathway are fused and expressed with these two receptor proteins. The two receptor proteins interact under the induction of GA, and the fusion signaling pathway is also induced by GA. The present invention selects the key protein LRP6 on the human Wnt pathway. After the fusion expression of GA receptor and LRP6, LRP6 will aggregate under the induction of GA, thereby promoting the production of β-catenin and opening the intracellular signaling pathway. After adding PAC inhibition, the production of β-catenin is reduced and the cell pathway is closed.

Description

technical field [0001] The invention relates to the field of biological method inventions, specifically, a method for regulating cell pathways by using plant hormone GA and small molecular substance PAC. Background technique [0002] Optogenetics is a new technology that combines light control technology with genetics for cell biology research, that is, expresses light-sensitive ion channel proteins on excitable target cells or organs, and activates them with light of corresponding wavelengths. Light-sensitive channels are used to fine-tune the physiological functions of cells, tissues, organs and animals. So far, many photoreceptors have been improved and used to fuse key proteins in signaling pathways to regulate and control many pathways in organisms. Such as receptor tyrosine kinase Ras-MAPK signaling pathway, PI3K-Akt signaling pathway and Wnt signaling pathway. At the gene level, the researchers used elements such as CRY2-CIB1, FKF1-GI, UVR8-COP1, and LOV proteins to ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85C12N5/10
CPCC12N15/85C12N2840/002
Inventor 左泽乘张峻川莫伟亮张力肖勇石翔张宝昌张文礼李苏迪
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