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A method for gene editing of sturgeon talen

A gene editing and sturgeon technology, applied in genetic engineering, biochemical equipment and methods, and other methods of inserting foreign genetic materials, can solve the problem of establishing gene editing technology without sturgeon

Active Publication Date: 2020-10-30
BEIJING FISHERIES RES INST +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, no gene-editing technology has been established in sturgeon so far

Method used

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  • A method for gene editing of sturgeon talen
  • A method for gene editing of sturgeon talen
  • A method for gene editing of sturgeon talen

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] A method for gene editing of sturgeon TALEN, comprising the steps of:

[0026] 1) Select sexually mature sterlet broodstock and temporarily raise them in an indoor culture system. 25 hours before the artificial breeding, inject the mixture of LHRHa and DOM to artificially induce labor, the injection dose is 10μg LHRHa+1mg DOM / kg body weight, inject once every 12 hours, the female sturgeon is injected twice continuously, and the male sturgeon is injected once Second-rate. When the broodstock has signs of ejaculation / ovulation, squeeze out the semen into a dry beaker and squeeze out the eggs into a glass dish. Use 0.3×Danieau buffer [17mM NaCl, 2mM KCl, 0.12mM MgSO 4 , 1.8mM Ca(NO 3 ) 2 , 1.5mM HEPES, pH 7.6] Dilute the semen 100 times for artificial insemination. The fertilization time is 1-2 minutes, and then the excess semen is washed away to obtain fertilized eggs of sterlet that can be used for microinjection. Place the fertilized eggs in a slightly covered 0.3...

Embodiment 2

[0042] The same target gene was selected, and according to the method in Example 1, micromanipulation of embryos was successfully carried out in A. schrenckii, A. gueldenstaedtii and A. baerii.

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Abstract

The invention discloses a sturgeon TALEN gene editing method. The sturgeon TALEN gene editing method comprises the following steps: mixing target gene plasmids with TALEN mRNA in allusion to target sites, and then injecting into micropyles of animal poles of 1-cell stage fertilized eggs of a sturgeon with the fertilization time being not longer than 20 minutes in a micromanipulation way, wherein after the target gene plasmids and the TALEN mRNA in allusion to the target sites are mixed according to a concentration ratio of 1:3, the plasmids with the concentration of 100 ng / [mu]L are introducedinto 1-cell stage fertilized embryos of a sterlet by a microinjection method, so that sterlet embryos with the survival rate of 74% and the exogenous target gene mutation rate as high as 93% can be obtained. By the sturgeon TALEN gene editing method, an accurate exogenous sturgeon gene editing technology is established for the first time, and powerful technical means for carrying out functional study on a sturgeon gene and genetic improvement on a cultured sturgeon species are provided.

Description

technical field [0001] The invention relates to the technical field of aquatic animal genetic breeding and functional gene verification research, and more specifically relates to a method for gene editing of sturgeon. Background technique [0002] Sturgeon is in the transition period between cartilaginous fish and bony fish in terms of biological system evolution, and has a polyploidized genome, which has important research value in the study of fish gene function and evolution. However, the existing reports on gene function research in sturgeon are limited to gene cloning and expression pattern analysis (Abdolahnejad et al., 2015. Dong et al., 2015. Fajkowska et al., 2016. Song et al., 2016. Yarmohammadi et al., 2017) . Sturgeons are large economic fish. Sturgeon is not only delicious in quality, comprehensive in nutrition, rich in a variety of essential amino acids and unsaturated fatty acids, but also an important material for caviar production (Simeanu et al., 2012). ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85C12N15/89
CPCC07K14/461C12N15/8509C12N15/89C12N2800/106C12N2800/80
Inventor 胡红霞陈戟胡炜朱华王巍朱作言
Owner BEIJING FISHERIES RES INST
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