SgRNA and application thereof

A targeted and basic sequence technology, applied in sgRNA and its application fields, to achieve high editing efficiency, knock down the level of PTBP1mRNA, and reduce the level of the effect

Active Publication Date: 2021-03-19
GUANGZHOU REFORGENE MEDICINE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] There is no research on targeted editing of

Method used

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  • SgRNA and application thereof
  • SgRNA and application thereof
  • SgRNA and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0061] 1. Design and construction of sgRNA vector

[0062] 1. sgRNA design.

[0063] The sgRNA targeting PTBP1 mRNA is designed, and its targeting domain sequence (reverse complement to the target sequence on the mRNA molecule) is 20-30nt in length.

[0064] The targeting domains of the designed sgRNAs are shown in Table 1 below. Figure 2-12 The target position of the designed sgRNA on PTBP1 mRNA is shown, wherein the box indicates the target site of the sgRNA, and the number in the box is the number of the sgRNA.

[0065] Among them, sgRNA 1-11 and sgRNA 32-50 designed by the present invention were used in the following plasmid transfection mouse N2A cell experiment. In the following experiment of plasmid transfection into human 293T cells, the sgRNA 1-89 designed by the present invention was used.

[0066] Among them, sgRNA 1-11 and sgRNA 32-50 target human and mouse homologous sequences.

[0067] sgRNA 1-20 and sgRNA 32-67 can target the CDS region of human PTBP1 mRNA....

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Abstract

The invention relates to sgRNA and application thereof, and belongs to the technical field of gene editing. The sgRNA sequence damages PTBP1 mRNA in a targeted manner, and the structural domain sequence of the sgRNA sequence is selected from any one of basic sequences shown in SEQ ID NO:1-31, or a truncated sequence which has the sequence length of more than or equal to 20nt and is obtained by truncating at least one basic group at the 3' end and/or the 5' end of the basic sequence, or an extended sequence which has the similarity of more than or equal to 90% with the basic sequence or the truncated sequence. With adoption of the sgRNA, the human PTBP1 mRNA can be destroyed in a targeted manner, the PTBP1 mRNA level can be effectively reduced, and the sgRNA is used for treating PTBP1 mRNAoverexpression related diseases or treating related diseases such as amyotrophic lateral sclerosis, Alzheimer's disease, Parkinson's disease and Huntington's disease by lowering the conventional PTBP1mRNA level, and has important application significance.

Description

technical field [0001] The invention relates to the technical field of gene editing, in particular to a sgRNA and its application. Background technique [0002] Neurodegenerative diseases are characterized by the progressive and irreversible loss of neurons from specific regions of the brain. The pattern of neuronal loss is selective, primarily affecting subcortical regions (basal nuclei) and the cerebral cortex, resulting in abnormal voluntary motor control, memory loss, and cognitive decline. Eventually lead to amyotrophic lateral sclerosis, Alzheimer's disease, Parkinson's disease and Huntington's disease. At present, there are still few therapeutic options for the effective treatment of these neurodegenerative diseases, and there is still an urgent clinical need for corresponding therapeutic drugs. [0003] A promising direction of current research is the use of regenerative techniques to regrow healthy and functional neural tissue in damaged areas. Various approaches...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12N15/85C12N9/22A61K48/00A61K38/46A61P25/28A61P25/16A61P25/14A61P25/00A61P21/00
CPCC12N15/113C12N15/85C12N9/22A61K48/005A61K38/465A61P25/28A61P25/16A61P25/14A61P25/00A61P21/00C12N2310/20C12N2800/107Y02A50/30
Inventor 梁峻彬王士民徐辉古博
Owner GUANGZHOU REFORGENE MEDICINE CO LTD
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