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Goat MSTN (myostatin) gene fixed-point modification system and application thereof

A site-directed modification, gene technology, applied in the field of genetic engineering, can solve the problems of small fragment base loss or insertion, affecting gene function, etc.

Active Publication Date: 2014-07-30
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

DSBs activate two conserved DNA repair pathways in eukaryotic cells: non-homologous end joining (NHEJ) can reconnect broken chromosomes, and the break site may introduce small fragments during the joining process Loss or insertion of bases, thereby affecting gene function or producing gene knockout effect

Method used

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  • Goat MSTN (myostatin) gene fixed-point modification system and application thereof
  • Goat MSTN (myostatin) gene fixed-point modification system and application thereof
  • Goat MSTN (myostatin) gene fixed-point modification system and application thereof

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Embodiment Construction

[0036] The present invention is described in further detail below in conjunction with accompanying drawing and specific implementation case, but not as the limitation of the present invention, and concrete operation process is carried out according to the following steps:

[0037] The first step is to design a specific sequence TALEN target according to the gene sequence to be knocked out;

[0038] The second step is to construct a TALEN plasmid according to the designed TALEN target;

[0039] Step 3: Transfect recipient cells with TALEN plasmids;

[0040] The fourth step is to screen, cultivate, and pick monoclonal cells after the transformation, expand the culture of monoclonal cells, identify the genomic mutation of the monoclonal cells, and select positive cell clones that meet the mutation requirements for subsequent experiments.

[0041] The experimental methods used in the following examples are conventional methods unless otherwise specified.

[0042] Materials, reag...

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Abstract

The invention discloses a goat MSTN (myostain) gene fixed-point modification system and an application method thereof. The goat MSTN gene fixed-point modification system can effectively identify and modify transcription activator like effector nucleases (TALENs) of a goat MSTN gene target sequence, the TALENs provided by the invention is composed of nucleases TALEN-R and TALEN-L which can respectively identify identification modules TALMEX-1F and TALMEX-1R; the nuclease TALEN-L is a protein coded by a nucleotide sequence shown in SEQ ID NO.2 or SEQ ID NO.3, and the nuclease TALEN-R is a protein coded by a nucleotide sequence shown in SEQ ID NO.4 or SEQ ID NO.5.The invention also discloses a method for obtaining a muted cell line by carrying out targeted modification on a goat cell MSTN gene by adopting the knock-out system. According to the goat MSTN gene fixed-point modification system, the goat MSTN gene is knocked out or modified for obtaining a new variety with a high high-quality meat factor, so that the goat MSTN gene fixed-point modification system has an important application value on mechanism study of an MSTN gene regulation and control mechanism.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a pair of transcription activator-like effector nucleases and their coding genes and their application, specifically, using a pair of transcription activator-like effector nucleases to modify goat MSTN gene at a fixed point. Background technique [0002] Myostatin (MSTN) widely exists in animal skeletal muscle and is a negative regulator of skeletal muscle growth. MSTN gene is expressed in skeletal muscle during animal embryonic development and adulthood. The cDNAs of MSTN genes in mice, rats, humans, cattle, pigs, sheep, chickens, turkeys, and zebrafish have been cloned and sequenced. The results showed that MSTN gene contains 3 exons, and the sequence of MSTN gene is highly conserved among different species. Knocking out the MSTN gene will lead to extensive hyperplasia and hypertrophy of animal muscle fibers, and a significant increase in skeletal muscle mass. Poi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12N15/63C12N1/15C12N1/19C12N1/21
Inventor 成勇于宝利
Owner YANGZHOU UNIV
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