Method for carrying out interspecies cross artificial insemination on frozen semen of paralichthys dentatus and paralichthys olivaceus
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A technology of frozen semen and flounder, which is applied in the field of marine biology, can solve the problems of interspecies hybrid sperm, eggs, asynchronous gonad development, etc., and achieve the effects of good repeatability, high recovery rate, and avoiding damage
Inactive Publication Date: 2013-10-16
INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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Problems solved by technology
However, flounder and summer flounder have different biological characteristics, and the gonad development is often asynchronous during the artificial breeding process. At
Method used
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Embodiment 1
[0019] Collect the sperm of flounder at the peak reproductive stage, that is, collect the sperm of flounder in Yantai Dongfang Marine Broodstock Breeding Workshop, and store them in a clean centrifuge tube. The viability of the microscope is higher than 90%, and they are placed on crushed ice and brought back to the laboratory for further analysis. cryopreservation;
[0020] Mix the fresh sperm collected above with antifreeze in crushed ice (0°C) at a volume ratio of 1:3, shake gently, mix well for 3 minutes, and distribute them into 2ml cryopreservation tubes, and then place them in a cooling tube. Equilibrate in the instrument at 0°C for 5 minutes; then cool down to -150°C at a cooling rate of -10°C / min, pack into cryopreservation boxes, place them in liquid nitrogen containers for long-term ultra-low temperature cryopreservation, and store 65 tubes of frozen semen;
[0021] Among them, the antifreeze is composed of antifreeze, antifreeze and diluent; the diluent has a mass ...
Embodiment 2
[0027] In May 2010, March 2011, and December 2012, the sperm of flounder was collected in Yantai Dongfang Ocean Broodstock Breeding Workshop, and the viability of the microscopic examination was higher than 90%. Place the fresh sperm on crushed ice and bring it back to the laboratory for cryopreservation, or directly store the fresh sperm collected above on site.
[0028] The fresh essence is mixed with antifreeze solution at a volume ratio of 1:3, shaken gently in crushed ice (0°C), mixed thoroughly for 3 minutes, and distributed into 2ml cryopreservation tubes, and then placed in a cooling device at 0°C Equilibrate for 5 minutes; then cool down to -150°C at a cooling rate of -10°C / min, pack into cryopreservation boxes, place in liquid nitrogen containers for long-term ultra-low temperature cryopreservation, and store 65 tubes of frozen semen;
[0029] Among them, the antifreeze is composed of antifreeze, antifreeze and diluent; the diluent has a mass concentration of 8g / L Na...
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Abstract
The invention belongs to the technical field of marine organisms, particularly a method for carrying out interspecies cross artificial insemination on an egg of paralichthys olivaceus and a semen of paralichthys dentatus, which is frozen for preservation. The semen of paralichthys dentatus, which is frozen for preservation at an ultralow temperature for a long time, is dissolved in water by two steps; after the semen of paralichthys dentatus is dissolved, diluent of which the volume is three times of that of the frozen semen of paralichthys dentatus is added into the frozen semen of paralichthys dentatus; after the frozen semen of paralichthys dentatus and the diluent are mixed, centrifugation is carried out for 5min; then supernate is removed to obtain frozen semen suspension with the same volume with the frozen semen of paralichthys dentatus before the frozen semen of paralichthys dentatus is diluted; the frozen semen and natural seawater are uniformly mixed according to the proportion of a volume ratio of 1:40 and the mixture is activated; after the mixture is activated, the egg of paralichthys olivaceus is injected and the mixture and the egg of paralichthys olivaceus are slightly and uniformly mixed to enable the egg of paralichthys olivaceus and the frozen semen of paralichthys dentatus to be sufficiently in contact; standing is carried out for 5 to 10min and egg washing is carried out to obtain a fertilized egg to carry out incubation. The invention establishes a cryopreservation technology for semen of paralichthys dentatus and a method for interspecies cross artificial insemination on the egg of paralichthys olivaceus, overcomes the difficulty of asynchrony of development of parental fishes of paralichthys dentatus and paralichthys olivaceus and provides a novel method and a novel idea for seawater fish cross breeding.
Description
technical field [0001] The invention belongs to the technical field of marine organisms, and specifically relates to an artificial insemination method for interspecies hybridization of flounder ova and cryopreserved flounder sperm. Background technique [0002] Cross-breeding is a breeding method that is relatively easy to implement and effective. It uses existing varieties or species to carry out crossbreeding, and selects a gene combination that meets people's needs from its offspring. Selecting geographically distant species as hybrid parents can increase the polymorphism of hybrid progeny, and it is easy to obtain a variety of types from their hybrid progeny, and it is relatively easy to obtain new types that meet the requirements. Flounder (Paralichthys olivaceus) and summer flounder (Paralichthys dentatus) are important marine fish species, which are distributed in the Pacific and Atlantic coasts respectively. Although they belong to the same genus, their geographical ...
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