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Goldfish gene editing technology and method for creating new goldfish variety by using goldfish gene editing technology

A technology of gene editing and new species, applied in the field of fish molecular design and breeding, can solve the problems of lack of fast and efficient systematic methods, achieve the effects of reducing background differences, avoiding off-target phenomena, and improving gene editing efficiency

Pending Publication Date: 2021-04-20
INST OF AQUATIC LIFE ACAD SINICA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, effective goldfish gene editing technology has not yet been established, and how to use gene editing technology to regulate the phenotype of goldfish and obtain offspring with a stable phenotype, there is no fast and efficient systematic method

Method used

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  • Goldfish gene editing technology and method for creating new goldfish variety by using goldfish gene editing technology
  • Goldfish gene editing technology and method for creating new goldfish variety by using goldfish gene editing technology
  • Goldfish gene editing technology and method for creating new goldfish variety by using goldfish gene editing technology

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Efficient breeding of pleated skirt Thai lion goldfish

[0031] S1. Seed cultivation: Raise freshly opened fry in a turnover box with a water temperature of 24-26°C and feed live Artemia 3 times a day. The turnover box is 60-100cm long, 40-60cm wide, 20-30cm high, and the water level is 15 -20cm is appropriate. Install a sponge filter on one side of the turnover box to filter, and place an oxygen increasing head next to it for oxygenation, and the air volume is appropriate. Clean the sponge filter once a week.

[0032] After the fry exceed 2cm in length, look down to select four-tailed fry, and then look sideways to remove the deformed and folded fry, and put the selected high-quality fry into a new container for rearing, with a density of about 2 fish per liter of water; the container is equipped with 7 watts small water pump, trickle to add water, the daily water intake is about 1 / 5 of the total water volume, and the three-layer turnover box is filtered b...

Embodiment 2

[0043] Example 2: Exploration of Gynogenetic Conditions

[0044] Two methods of gynogenesis were established, the proportion of normal fish produced by the two methods was compared, and the one with a higher rate of homozygosity was selected as the best gynogenesis condition for Thai lion goldfish.

[0045] 1. Meiotic gynogenesis

[0046] Take carp sperm to inactivate, mix inactivated sperm with pleated skirt Thai lion goldfish eggs, and then perform cold shock treatment. The cold shock treatment method is to develop fertilized eggs at a constant temperature of 24°C for 3-7 minutes, and then place them at 0°C Cold shock was carried out in ice water for 2 minutes to inhibit the discharge of the second polar body, so that the two gametes merged into one to form a tetraploid. After the cold shock treatment, the fertilized eggs hatched into meiotic Thai lion goldfish with dragon eye traits.

[0047] 2. Mitotic Gynogenesis

[0048] Take carp sperm to inactivate, mix the inactiva...

Embodiment 3

[0053] Example 3: Creation of Dragon Eye Pleated Skirt Thai Lion Goldfish

[0054] The pleated skirt Thai lion is one of the most important ornamental species of side-viewing goldfish. The dragon eye character is named for its protruding eyes, which are similar to the eyes of dragons in myths and legends. The dragon eye character plays an important role in overlooking goldfish. It is the dream of many goldfish farms and goldfish lovers to transfer the dragon eye trait from traditional ornamental fish to Thai lion. However, the traditional method of continuous hybridization and purification cannot achieve the transfer of a single dragon eye trait. After the dragon eye trait is transferred to the pleated skirt Thai lion goldfish through hybridization, it will be accompanied by changes in other important ornamental traits of the pleated skirt Thai lion goldfish. For example, the angle between the tail becomes smaller, there are no folds, and the body shape is no longer thick, thu...

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Abstract

The invention relates to a fish molecular design breeding technology, in particular to a goldfish gene editing technology and a method for creating a new goldfish variety by using the gene editing technology. The method comprises steps of knockout target design, microinjection, fertilization incubation, gynogenesis and high-temperature sex reversal. According to the invention, a CRISPR / Cas9 gene editing technology is established on goldfish, so that a target phenotype is generated on F0 generation goldfish, an efficient gynogenesis condition of pleated Thai lion goldfish is determined through experiments, homozygotes are obtained in F1 generation, and a large number of groups are expanded in F2 generation through temperature-induced reversion. Through a molecular breeding means, transfer of a single target character is rapidly achieved, the goldfish gene editing technology and the method can be used for creating a new goldfish variety, provides technical support for molecular design and breeding of goldfish, and is beneficial to promote development of goldfish molecular breeding technology.

Description

technical field [0001] The invention relates to fish molecular design and breeding technology, in particular to a goldfish gene editing technology and a method for creating new goldfish varieties using the gene editing technology. [0002] technical background [0003] Goldfish originated from the tetraploid crucian carp in my country. It is an ornamental fish formed after hundreds of years of artificial selection, and it can be called the quintessence of our country. In the history of goldfish development, due to mutations, goldfish body color, body shape and almost all external organs have undergone obvious changes. Due to artificial selection, these mutations have been retained, and these mutations have been hybridized and recombined to form multiple compound variant goldfish; Today, there are more than 300 species of goldfish. [0004] Traditional goldfish are kept in fish pots, and the breeding of species is mainly based on looking down. The popularity of fish tanks ha...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/89A01K67/027
Inventor 余鹏桂建芳周莉汪洋李志李亮亮杨小丽
Owner INST OF AQUATIC LIFE ACAD SINICA
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