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Method for inducing misgurnus anguillicaudatus gynogenesis tetraploid fries by using heterogenenos sperms

A technique for gynogenesis and tetraploidy, applied in the field of loach gynogenetic tetraploid fry induced by heterologous sperm, achieving the effects of advanced technology, simplified identification problems, and efficient methods

Active Publication Date: 2015-05-13
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is no report on the use of bream sperm to induce gynogenesis and chromosome manipulation in the eggs of natural tetraploid loach

Method used

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  • Method for inducing misgurnus anguillicaudatus gynogenesis tetraploid fries by using heterogenenos sperms
  • Method for inducing misgurnus anguillicaudatus gynogenesis tetraploid fries by using heterogenenos sperms
  • Method for inducing misgurnus anguillicaudatus gynogenesis tetraploid fries by using heterogenenos sperms

Examples

Experimental program
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Effect test

Embodiment 1

[0043] A method for inducing tetraploid fry of loach gynogenetic tetraploid fish induced by heterologous sperm, the steps are as follows:

[0044] A. Selection of parents and artificial induced labor

[0045] Select tetraploid female loach with normal body shape, no injury in appearance, strong physique, swollen and elastic abdomen, and obvious outline of ovary. Male bream requires obvious pectoral fin pearls, rough feeling, and white semen flowing out from the genital opening when the abdomen is lightly pressed.

[0046] B. Genetic inactivation of sperm

[0047] The sperm of the group head bream was collected, and those whose vigor was above 90% were selected for ultraviolet inactivation by microscopic examination. Take 1ml of the group head bream semen, dilute it with Hank's solution at a volume ratio of 1:4, and place it in a pre-cooled dry petri dish (dia. 75mm), the semen thickness is about 0.1 or 0.15 or 0.2mm, and then placed under two 15W ultraviolet lamps for treatmen...

Embodiment 2

[0059] A method for inducing tetraploid fry of loach gynogenetic tetraploid fish induced by heterologous sperm, the steps are as follows:

[0060] A. Selection of parents and artificial induced labor

[0061] Select tetraploid female loach with normal body shape, no injury in appearance, strong physique, swollen and elastic abdomen, and obvious outline of ovaries. Male bream requires obvious pectoral fin pearls, rough feeling, and white semen flowing out from the genital opening when the abdomen is lightly pressed.

[0062] B. Genetic inactivation of sperm

[0063] Collect bream sperm, microscopically select those with vigor above 90% for ultraviolet inactivation, take 1ml of the above-mentioned treated bream semen, dilute it with Hank's solution at a volume ratio of 1:4, and place it in a pre-cooled dry In a Petri dish (75mm in diameter), the thickness of the semen is about 0.15mm, and then placed under two 15W ultraviolet lamps for treatment. The distance between the lamp...

Embodiment 3

[0087] A method for identification of loach gynogenetic tetraploid fry, the steps are as follows:

[0088] Tetraploid fry were identified using one of the following two methods.

[0089] A. Analysis and identification of gynogenetic tetraploid fry by flow cytometry and chromosome counting method;

[0090] Fertilized eggs were hatched after cleavage stage, blastula stage, gastrula stage, neurula stage, yolk embolus stage, Kirschner blastula stage, tail tooth embryo stage and heartbeat stage, and the 15-day-old Fish fry were anesthetized with MS-222, and some tissues were cut into pieces from each individual, and the rest were preserved in 95% absolute ethanol for future use. These partial tissue sections were crushed in a tissue homogenizer equipped with 300 μl PBS, filtered through a 200-mesh nylon mesh, the cell suspension was collected, centrifuged at 800r / min for 6min, the supernatant was discarded, and stained with DAPI staining solution for 5min After testing on the mac...

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Abstract

The invention discloses a method for inducing misgurnus anguillicaudatus gynogenesis tetraploid fries by using heterogenenos sperms. The method comprises the following steps: selecting parents, performing genetic inactivation on sperms, inducing gynogenesis tetraploid fries, and performing cold shock treatment to obtain gynogenesis tetraploid fries. The highest fertility rate, hatching rate and survival rate of the gynogenesis tetraploid misgurnus anguillicaudatus can reach 76.99+ / -2.51%, 54.54+ / -1.87% and 17.9+ / -0.65% respectively. The method disclosed by the invention has the characteristics of advancement, high efficiency, practicability and reliability, and has an important application value and a wide application prospect in the breeding of misgurnus anguillicaudatus.

Description

technical field [0001] The invention belongs to the field of loach gynogenesis induced by heterologous sperm, in particular to a method for heterologous sperm inducing loach gynogenetic tetraploid fry. Background technique [0002] Gynogenesis is an important reproductive mode in fish parthenogenesis, which uses genetically inactivated sperm to activate the development of mature eggs. After the sperm enters the egg, it does not undergo fusion of the male nucleus and the egg nucleus, but only activates the egg. Initiate the role of development, and then induce gynodiploidy to obtain gene homozygous fish. Artificial gynogenesis is of great significance in the rapid establishment of fish pure lines, sex determination and artificial control of sex, and the establishment of gene maps. The purity of one artificial induction of gynogenesis is equivalent to the result of 14 generations of full-sib mating and breeding. Any fish that undergoes two consecutive gynogenesis can be used ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01K61/00
CPCY02A40/81
Inventor 周小云于永耀王卫民
Owner HUAZHONG AGRI UNIV
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