Method for inducing protein interaction and gene expression by pesticide

A technology for inducing protein and gene expression, applied in the direction of microorganism-based methods, chemical instruments and methods, biochemical equipment and methods, etc., to achieve the effect of increasing usage

Pending Publication Date: 2021-08-10

NANTONG UNIVERSITY

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AI-Extracted Technical Summary

Problems solved by technology

[0006] The present invention provides a method for pesticides to induce protein interaction and gene expression, which solves the limitations of existing compounds that induce protein interaction as gene expr...

Method used

Plasmid 1 sequence is as shown in SEQ ID NO.1, and plasmid 2 sequences are as shown in SEQ ID NO.2, and plasmid 3 sequences are as shown in SEQID NO.3, and plasmid 4 sequences are as shown in SEQ ID NO.4, The sequence of plasmid 5 is shown in SEQ ID NO.5, the sequence of T7 N is shown in SEQ ID NO.6, the sequence of ABI1-CP is shown in SEQ ID NO.7, the sequence of PYR1MANDI is shown in SEQ ID NO.8, and the sequence of T7C As shown in SEQ ID NO.9, GFP10 sequence as shown in SEQ ID NO.10, GFP11 sequence as shown in SEQ ID NO.11, GFP1-9 sequence as shown in SEQ ID NO.12, T7 promoter_lacO sequ...

Abstract

The invention discloses a method for inducing protein interaction and gene expression by a pesticide, the pesticide is mandipropamid, the protein interaction is induced as green fluorescent protein three-fragment complementation, and the gene expression is induced as green fluorescent protein coding gene expression; the provided ABI-CP is a topological structure variant using mandipropamid to control protein interaction, and the limitation of the existing method on the relative position of the fusion protein space is broken through; according to the method for regulating and controlling the gene expression by using mandipropamid, the limitation of the existing compound for inducing the gene expression is broken through due to the characteristics of the pesticide compound. The limitations include high cost, incapability of being massively used in the environment, non-physiologically neutral (capable of being rapidly metabolized), difficulty in entering cells to take effect and the like.

Application Domain

Microorganism based processesPlant peptides +3

Technology Topic

EGFP proteinPesticide +6

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Examples

Experimental program(2)

Example Embodiment

[0046] Example 1:

[0047] A pesticide-induced protein interaction and induction gene expression, butachalactamine induces complementary green fluorescent protein three fragments, steps:

[0048] Step 1: Construct a plasmid 2 on the basis of the plasmid 1, ABI-CP amino end fusion GFP10, PYR1 MANDI Carboxy end fusion GFP11.

[0049] Step 2: The plasmid 3 was constructed, controlled by T7 promoter _Laco in E. coli to induce GFP1-9 fragments to match the IPTG induction system of E. coli BL21 (DE3) strain.

[0050] Step 3: Transform plasmids 2 and 3 Transform Escherichia coli BL21 (DE3) strain.

[0051] Step 4, the expression product of the detection of plasmids 2 and 3 is induced by bisyl bacillus in E. coli, and positive cloning rhubine in the background of E. coli BL21 (DE3) strain, to be OD 600 To 0.40-0.60, IPTG (isopropylthiolanside) induced by 1 mM was added to the bacterial liquid induced GFP1-9 fragment expression, and the final concentration gradient was added to bisyltysyl amine.

[0052] Step 5: After 3-4 hours, use the following parameters, excitation light 488 nm, and transmit light 510nm to detect fluorescence intensity of GFP1-9, GFP10, and GFP11 three segments as GFP, while determine OD 600 Used in homogenic bacterial liquid culture concentrations, showing that bisthynthynthynacaine is highly efficient induced green fluorescent protein three fragments complementary.

Example Embodiment

[0053] Example 2

[0054] Dikaronyltysin induces green fluorescent protein coding gene expression:

[0055] The first step: the plasmid 4 is constructed, and the Split T7 RNA polymerase and PYR1 are made in E. coli. MANDI And ABI-CP fusion expression, respectively;

[0056] Step 2: The plasmid 5 is constructed, and the GFP coding gene expression is used to detect the activity of the T7 RNA polymerase in E. coli by T7 promoter.

[0057] Step 3: Plasmids 4 and 5 transform Escherichia coli TOP10 strains. Note that strains with endogenous T7 RNA polymerase cannot be used, such as BL21 (DE3) strain;

[0058] Step 4: Detecting the expression product of plasmids 4 and 5 induced by bisyl bacilline in E. coli: Take the positive cloning rhubine on the background of Escherichia coli TOP10 strain, to be OD 600 When reaching 0.40-0.60, the final concentration gradient was added to the bacterial solution of a hypertysyl esterine of 0.1 μm, 1 μm, 10 μm, 100 μm, 200 μm, and 400 μm, 3-4 hours, using the following parameters, excited light 488 nm, and emit 510 nm detection GFP expression effect, while determining OD 600 Used in homogenic acid culture concentration, showing bisthynthynthynacalamine induces green fluorescent protein encoding gene expression.

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Method for inducing protein interaction and gene expression by pesticide

A technology for inducing protein and gene expression, applied in the direction of microorganism-based methods, chemical instruments and methods, biochemical equipment and methods, etc., to achieve the effect of increasing usage

AI-Extracted Technical Summary

Problems solved by technology

Method used

Abstract

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Examples

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Example Embodiment

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Description & Claims & Application Information

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