Method for creating dragon-eye pleated Thai-cultured lion-head goldfish

A technology of pleated skirts with dragon eyes and Thai lion goldfish is applied in the field of molecular design and breeding of fish, which can solve the problems of losing the ornamental value of pleated skirts and the body shape is no longer thick.

Pending Publication Date: 2021-05-18
INST OF AQUATIC LIFE ACAD SINICA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, after years of attempts, the traditional method of continuous hybridization and purification cannot achieve the transfer of single-purpose traits. The dragon eye trait is transferred to the pleated skirt Thai lion goldfish through hybridization, which will be accompanied by other important ornamental traits of the pleated skirt Thai lion goldfish. For example, the angle between the tail becomes smaller, there are no pleats, and the body shape is no longer thick, thus losing the original ornamental value of the pleated skirt Thai lion

Method used

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  • Method for creating dragon-eye pleated Thai-cultured lion-head goldfish
  • Method for creating dragon-eye pleated Thai-cultured lion-head goldfish
  • Method for creating dragon-eye pleated Thai-cultured lion-head goldfish

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1: Efficient breeding of pleated skirt Thai lion goldfish

[0033] S1. Seed cultivation: Raise freshly opened fry in a turnover box with a water temperature of 23-25°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.

[0034] 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 The 7-watt small water pump adds water by trickle, and the daily water intake is about 1 / 5 of the total water body. The three-layer turnover box is filtered ...

Embodiment 2

[0044] Example 2: Exploration of Gynogenetic Conditions

[0045] 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.

[0046] 1. Meiotic gynogenesis

[0047] 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.

[0048] 2. Mitotic Gynogenesis

[0049] Take carp sperm to inactivate, mix the inactiva...

Embodiment 3

[0054] Example 3: Creation of Dragon Eye Pleated Skirt Thai Lion Goldfish

[0055] S1. Target design: Sequence the gene of the pleated skirt Thai lion goldfish to obtain the DNA sequence and amino acid sequence of the gene that regulates the traits of the dragon eye, and use the wildcard GG[G / A]? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? GG or CC? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? [T / C] CC designs a suitable knockout target on the exon of the DNA sequence of the gene that regulates the traits of dragon eyes, builds a local genome database after obtaining the target, and uses blast software to search and verify the uniqueness of the target After determining the uniqueness, the target sequence is transcribed into gRNA in vitro; "?" in the wildcard character represents an arbitrary base, "[]" represents a match with any base in [], and " / " represents "or".

[0056] The gene regulating the traits of dragon eyes is lrp2a, the low-density lipoprotein 2 gen...

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Abstract

The invention relates to molecular design breeding technologies for fishes and particularly relates to a method for creating dragon-eye pleated Thai-cultured lion-head goldfish. The method comprises the steps of target point designing, microinjection, and fertilization and incubation. According to the method, a low-density lipoprotein 2 gene lrp2a is knocked out based on a gene editing technology of CRISPR/Cas9, then, goldfish of F0 generates a dragon eye phenotype, Efficient gynogenesis conditions and high-temperature group enlarging conditions for pleated Thai-cultured lion-head goldfish are established through experiments, and F1 obtains dragon-eye pleated Thai-cultured lion-head goldfish homozygotes; and through temperature-induced reversion, F2 is subjected to mass group enlarging. According to the method, by means of molecular breeding, transferring of a single target trait is achieved rapidly, a variety with a high ornamental value, i.e., the dragon-eye pleated Thai-cultured lion-head goldfish is stably and rapidly created, and thus, the method has a broad application prospect and a market value.

Description

technical field [0001] The invention relates to fish molecular design breeding technology, in particular to a creation method of dragon eye pleated skirt Thai lion goldfish. [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, the body color, body shape and almost all external organs of goldfish have undergone obvious changes. Under the action of artificial selection, these mutations have been retained, and these mutations have been crossed and recombined to form multiple compound variations. goldfish; today, there are more than 300 species of goldfish. Among the multiple mutant phenotypes of goldfish, dragon eye is an important ornamental trait. The character of dragon eyes is named because of its protruding eyes, which are simila...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/89C12N9/22C12N15/12A01K67/027A01K61/10
CPCA01K67/0276A01K2217/075A01K2227/40A01K2267/02A01K61/10C07K14/775C12N9/22C12N15/89Y02A40/81
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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