Transcription activator like effector-functional group-estrogen receptor function protein and application thereof

A technology of transcription activator and estrogen receptor, applied in application, genetic engineering, plant genetic improvement, etc., can solve the problem that there is no way to control it accordingly

Inactive Publication Date: 2014-05-07
BEIJING VIEWSOLIDBIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the whole process, as long as the nucleic acid is introduced into the cell, there is no way to control it accordingly. In many experiments, such as nuclease experiments or recombinase experiments, in many cases, it is necessary to perform the test in a specific tissue at a specific time. Express the TALE fusion protein in a specific tissue and let it play a role, so as to show a specific phenotype in a specific tissue
These experiments, the current TALE technology has no way to achieve such an effect

Method used

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  • Transcription activator like effector-functional group-estrogen receptor function protein and application thereof
  • Transcription activator like effector-functional group-estrogen receptor function protein and application thereof
  • Transcription activator like effector-functional group-estrogen receptor function protein and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] The principle of embodiment 1 is as follows figure 1 As shown, the experimental results are as figure 2 and image 3 shown. The experimental process is as follows: First, in order to establish a detection system, the sequence containing the TALE binding site is integrated into the genome of the target cell through the method of virus infection. This is done for the convenience of operation. In practical application, On the premise of knowing the sequence of the target gene, a sequence in the target gene can be selected, and a TALE that binds to the sequence can be designed and synthesized according to the sequence. In this sequence, the TALE binding site is followed by a micro-promoter, and the expression level of the downstream reporter gene is very low in the absence of activation domain (ie, transcriptional activator (Activation Domain, AD)) stimulation. Then we did two experiments in parallel. In the first step, the plasmid containing ER-TALEa (that is, the fusi...

Embodiment 2

[0035] The experimental process of Example 2 is similar to that of Example 1, except that the AD protein was replaced by nuclease, and we selected a gene on the genome for the experiment (there is no need to integrate the target sequence into the genome by means of a virus). See the experimental results Figure 4 . Depend on Figure 4 It can be seen that both TALE-ER-nuclease and TALE-nuclease-ER can control their behavior of entering the nucleus through 4-hydroxy-tamoxifen, while TALE-nuclease-ER is more controllable. it is good.

Embodiment 3

[0036] The experimental process of Example 3 is similar to that of Example 1, except that the AD protein is replaced with a repressor protein (repressor), similar to Example 2, and a gene on the genome is selected for the experiment. See the experimental results Figure 5 . Depend on Figure 5 It can be seen that both TALE-ER-repressor and TALE-repressor-ER can control their behavior of entering the nucleus through 4-hydroxy-tamoxifen, while TALE-repressor-ER is more controllable. it is good.

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Abstract

The invention relates to a transcription activator like effector (TALE)-functional group-estrogen receptor (ER) function protein and an application thereof, wherein the TALE-functional group-ER function protein is formed by sequentially linking TALE, ER and a functional group or sequentially linking TALE, a functional group and ER, wherein the functional group is a combination comprising one or more than two materials selected from activation domain (AD), nuclease, methylase, demethylase and recombinase. During practical application, tamoxifen (TAM) or 4-OH-tamoxifen (4-OH-TAM) are introduced into cytoplasm so as to control the function protein to enter nucleus at the specific time.

Description

technical field [0001] The present invention relates to a "transcription activator like effector (Transcription Activator Like Effectors, TALE)-functional group-estrogen receptor (Estrogen Receptor, ER)" functional protein and the application of the above-mentioned functional protein, in particular to the above-mentioned functional protein The application of controlled access to the nucleus to manipulate specific genes. Background technique [0002] In the post-genome era, we urgently need new bioengineering technologies such as efficient gene manipulation and synthesis. For example, we often need to suppress or silence the expression of a gene. Traditional gene knockout (Gene Knockout) technology relies on homologous recombination that occurs naturally in cells, and its efficiency is very low, usually 10 -6 level; RNAi technology is simple and easy to implement, but it is difficult to obtain a 100% inhibitory effect. In addition to suppressing or silencing specific genes...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/00
Inventor 庄峰锋李娟
Owner BEIJING VIEWSOLIDBIOTECH
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