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Method for inhibiting HIV virus replication by targeting reverse transcription primer binding site

A target and inhibitor technology, applied in the field of basic research

Active Publication Date: 2022-06-24
TONGJI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, whether the zinc finger protein has any effect on the HIV virus and its primer-binding sequence PBS-Lys, and how the mechanism of action has not been reported

Method used

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  • Method for inhibiting HIV virus replication by targeting reverse transcription primer binding site
  • Method for inhibiting HIV virus replication by targeting reverse transcription primer binding site
  • Method for inhibiting HIV virus replication by targeting reverse transcription primer binding site

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1: Binding Zinc Finger Proteins The high throughput sequencing database identified ZNF417 and ZNF587 as potential PBS-Lys binding zinc finger proteins.

[0050] 1. Objective: To identify zinc finger proteins that bind HIV target PBS-Lys

[0051] 2. Method:

[0052] 1. By comparing the high-throughput zinc finger protein ChIPseq data of GSE78099 and GSE76496 in the GEO database with the PBS-Lys site, the candidate zinc finger proteins were screened out.

[0053] 2. By inserting the PBS-Lys site before the reporter gene vector promoter, and expressing it together with the zinc finger proteins ZNF417 and ZNF587 expression plasmids, the activity of the reporter gene was detected to verify the binding ability of ZNF417 and ZNF587 to PBS-Lys.

[0054] 3. The inhibitory effect of ZNF417 and ZNF587 on the PBS-Lys site was verified by performing ChIP experiments and high-throughput sequencing after HEK293T cells overexpressed GFP-tag ZNF417 and ZNF587.

[0055] 3. Resu...

Embodiment 2

[0057] Example 2: HIV pseudovirus infection experiment to verify the inhibitory effect of ZNF417 and ZNF587 on HIV

[0058] 1. Objective: To verify the ability of ZNF417 and ZNF587 to inhibit HIV

[0059] 2. Method:

[0060] 1. The luciferase reporter gene is driven by the LTR of HIV (similar to the promoter) and co-expressed with ZNF417 and ZNF587 to verify the ability of ZNF417 and ZNF587 to inhibit HIV transcription.

[0061] 2. By designing HIV pseudovirus (here using HIV full-length virus pNL4.3 and backbone virus pSicoR to express RFP, the RFP expression is used to indicate HIV virus infection efficiency) overexpression in ZNF417 and ZNF587 or gene knockout based on CRSIPRcas9 system The infected cells were infected by flow cytometry to determine whether ZNF417 and ZNF587 could inhibit HIV infection.

[0062] 3. By knocking out ZNF417 and ZNF587 in human CD4+ T cells and infecting HIV pseudovirus, simulating the role of ZNF417 and ZNF587 in HIV attacking host cells in ...

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Abstract

The invention belongs to the field of basic research, and in particular relates to a method for targeting the binding site of a reverse transcription primer to inhibit HIV virus replication. Firstly, a method for screening out HIV inhibitors from the database is disclosed, including: S1: extracting the recorded data files of zinc finger protein ChIP binding peaks from the database; S2: sorting out the HIV target sequence, and using the MEME tool to obtain the target motif Motif; Obtain the position information of the HIV target contained in the human genome; S3: Compare the ChIPseq binding peak file of the zinc finger protein with the position information file of the HIV target on the human genome, and obtain the potential HIV target inhibition zinc finger protein. The invention provides a method for finding HIV restriction factors, finds two restriction zinc finger proteins ZNF417 and ZNF587 of HIV, and fills in the blank in the field of human body's natural immunity to HIV.

Description

technical field [0001] The invention belongs to the field of basic research, and in particular relates to a method for targeting a reverse transcription primer binding site to inhibit HIV virus replication. [0002] technical background [0003] Retroviruses have long been attached to humans and other higher animals, co-exist and evolve with them, and are closely related to various biological phenomena. Retroviruses can be divided into endogenous and exogenous according to their relationship with the host. Endogenous retroviruses (ERVs) originate from the invasion of ancient exogenous retroviruses into mammalian germ cells, and gradually accumulate as part of the genome during long-term evolution. About 10% of the sequences in the mouse and human genomes belong to ERVs, and most of these ERVs are in a state of transcriptional silencing to maintain genome stability. Compared with ERVs, exogenous retroviruses are more pathogenic. For example, the well-known retrovirus human ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G16B20/30G16B35/20G16B5/00G16B40/00
CPCG16B20/30G16B35/20G16B5/00G16B40/00
Inventor 杨鹏王译萱杨博房璐
Owner TONGJI UNIV
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