A method for rapidly breaking diapause in indoor cultured arthropod eggs and improving hatching rate

Abstract

This invention discloses a method for rapidly removing diapause from dormant brine shrimp eggs cultured indoors and improving the hatching rate, relating to the field of brine shrimp culture using natural aquatic feed. The method includes the following steps: A. Collection and preservation of dormant brine shrimp eggs; B. Pretreatment of dormant brine shrimp eggs; C. Incubation under constant light. This method can shorten the diapause removal time for indoor-cultured brine shrimp eggs to less than 20 days and increase the hatching rate to 92.5%.

Description

Technical Field

[0001] This invention relates to the field of natural aquatic bait brine shrimp farming technology, and more specifically to a method for rapidly de-diapause dormant brine shrimp eggs and improve the hatching rate in indoor farming. Background Technology

[0002] Artemia, also known as brine shrimp, is a small, halophilic crustacean widely distributed in inland salt lakes and coastal salt fields. Artemia eggs and nauplius larvae are widely used in aquaculture as important high-quality protein feed. Furthermore, the unique biological characteristics of Artemia make it an excellent experimental material for studying crustacean embryonic development and a commonly used model organism for researching stress resistance mechanisms in aquatic organisms.

[0003] Artemia have two reproductive methods: oviparous and ovoviviparous. When environmental conditions are suitable, Artemia tend to adopt ovoviviparous mode, that is, directly producing nauplius larvae; while under harsh environmental conditions, oviparous mode is the main mode, producing dormant eggs with complex shells in a diapause state.

[0004] Artemia dormant eggs possess unique biological characteristics and special physiological and biochemical features. Their development is arrested, cell division stops, enzyme activity decreases, and metabolic activity is inhibited. They can be preserved for a long time and will hatch again when the external environment is suitable. However, even in the most suitable environment, the hatching rate of dormant eggs is very low. Only when stimulated by certain specific abiotic signals can this diapause state be terminated and normal physiological metabolism be restored.

[0005] Currently, there are many researches on methods and mechanisms for breaking dormancy in Artemia salina eggs, such as hydrogen peroxide treatment, ultraviolet light irradiation, long-term low-temperature treatment, or the addition of some chemical reagents, to break dormancy and promote the release of dormancy. However, these methods are not very effective and require a long time, often resulting in low hatching rates. Therefore, providing a rapid method for breaking dormancy and improving hatching rates is a problem that urgently needs to be solved by those skilled in the art. Summary of the Invention

[0006] In view of this, the present invention provides a method for rapidly dediapausing dormant brine shrimp eggs in indoor culture and improving the hatching rate. This method can rapidly dediapause brine shrimp eggs and improve the hatching rate.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] A method for rapidly breaking diapause in indoor cultured brine shrimp eggs and improving hatching rate includes the following steps:

[0009] A. Collection and preservation of dormant brine shrimp eggs

[0010] After cleaning, filtering, and drying the collected dormant brine shrimp eggs, they were stored in a -20℃ refrigerator for one week for low-temperature treatment.

[0011] B. Pretreatment of Artemia dormant eggs

[0012] First, the dormant eggs of Artemia after low-temperature treatment were placed in an incubator for the first light-proof and dark treatment (light intensity of 0 Lx) for 5 to 9 days; then, the dormant eggs of Artemia were transferred to an incubator equipped with UVB lamps for UVB irradiation treatment for 1 to 2 days; then the dormant eggs of Artemia were transferred back to the incubator for a second light-proof and dark treatment for 2 days.

[0013] C. Incubation under constant light

[0014] Formulating with added Fe 2+ Mg 2+ Artemia dormant eggs were added to the saline solution containing GnRH-a and incubated in a constant temperature incubator under constant light with a light intensity of 2000 Lx. The hatching rate was recorded after 1-2 days to determine the effect of removing diapause.

[0015] Preferably, the dormant eggs of Artemia in step A are dormant eggs produced by laboratory-cultured Artemia.

[0016] Preferably, the incubator in step B is a GTOP-310B2, BZ-GTOP-300Y, or NB-KRG-400A type intelligent light incubator.

[0017] Preferably, the initial light-avoidance dark treatment time in step B is 7 days, and the second light-avoidance dark treatment time is 2 days.

[0018] Preferably, the UVB irradiation treatment in step B is performed using a parallel ultraviolet beam, and the UVB irradiation equipment used is a GPH873T5L / 40W or Q-Lab / UVB-313EL type ultraviolet lamp.

[0019] More preferably, the total UVB irradiation dose in step B is 1.00 kJ / cm². 2 ~2.00kJ / cm 2 The irradiation dose was measured using a UVB radiometer, and the average UVB irradiation intensity was adjusted to 11.60 μW / cm² by adjusting the distance from the UV lamp. 2 According to the formula: UVB irradiation dose (mJ / cm²) 2 = Average light intensity (μW / cm) 2 Calculated by multiplying the irradiation time (s).

[0020] Preferably, the amount of dormant Artemia eggs added to the brine in step C is 1.0 g / L to 3.0 g / L.

[0021] Preferably, the brine preparation process in step C is as follows: dilute the aquaculture-specific sea salt with pure water to a concentration of 15‰ to 20‰ by mass-volume ratio; then add 5‰ to 15‰ Fe. 2+ and 5‰ to 15‰ Mg 2+ The final salt concentration was measured using a handheld ATC salinity refractometer to maintain the pH of the saline solution at 8.0–9.0; then GnRH-a was added to bring the final concentration to 0.5–1.5 mg / L.

[0022] More preferably, GnRH-a is added to the saline solution in step C to achieve a final concentration of 1.5 mg / L.

[0023] Preferably, the incubation temperature for both steps B and C is 26℃~30℃.

[0024] More preferably, the incubation temperature for both steps B and C is 28°C.

[0025] Beneficial effects:

[0026] 1. This invention provides a method for rapidly breaking diapause in indoor cultured artemisia eggs and improving hatching rates. Through the synergistic effects of low temperature, initial dark treatment, UVB irradiation, and secondary dark treatment, the method breaks the response of artemisia eggs to extreme environments. Changes in temperature and photoperiod promote the breaking of diapause in artemisia eggs. Combined with the stimulating effect of appropriate amounts of GnRH-a in the hatching brine, this method greatly promotes the metabolism of artemisia eggs, stimulating them to terminate diapause. Furthermore, appropriate amounts of Fe... 2+ and Mg 2+ This can induce the production of heme in the eggshell of Artemia salina, increasing the oxygen capture capacity of the eggs and improving the hatching rate. At the same time, low doses of UVB have a certain bactericidal effect, which can reduce the microorganisms carried by the Artemia salina eggs, reduce the competitive pressure for survival, and improve the hatching rate.

[0027] 2. The present invention provides a method for rapidly removing diapause from dormant Artemia eggs raised indoors and increasing the hatching rate. Through a series of stimuli and incubating Artemia under suitable light and temperature, the diapause time can be shortened to less than 20 days and the hatching rate can be increased to 92.5%. Detailed Implementation

[0028] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0029] This invention discloses a method for rapidly removing diapause from dormant brine shrimp eggs and improving hatching rate in indoor culture. Unless otherwise specified, the experimental reagents used in the following embodiments are all conventional biochemical reagents; the experimental instruments and methods described are all conventional laboratory instruments and methods; and the experimental terminology used has a generally accepted meaning in the field of this invention. All raw materials used are commercially available, and their sources are not specifically limited. Unless otherwise specifically mentioned, the methods involved are conventional methods and will not be described in detail here.

[0030] The dormant Artemia eggs used in Examples 1-4 were all parthenogenetic Artemia eggs.

[0031] Example 1

[0032] A method for rapidly dediapausing dormant brine shrimp eggs and increasing hatching rate includes the following steps:

[0033] A. Collection and preservation of dormant brine shrimp eggs

[0034] The collected Artemia dormant eggs were washed with tap water, filtered, and dried, and then stored in a -20℃ refrigerator for one week for low-temperature treatment; the Artemia dormant eggs were dormant eggs produced by artificially bred Artemia indoors.

[0035] B. Pretreatment of Artemia dormant eggs

[0036] First, place the dormant Artemia eggs in an incubator for an initial dark treatment for 5–9 days (i.e., light intensity of 0 Lx); then, transfer the dormant Artemia eggs to an incubator equipped with a UVB lamp for UVB irradiation treatment for 1–2 days; finally, transfer the dormant Artemia eggs back to the incubator for a second dark treatment for 2 days.

[0037] The incubator is a GTOP-310B2, BZ-GTOP-300Y, or NB-KRG-400A type intelligent light incubator.

[0038] The UVB irradiation treatment is performed using a parallel ultraviolet beam, and the UVB irradiation equipment used is a GPH873T5L / 40W or Q-Lab / UVB-313EL ultraviolet lamp.

[0039] The total UVB irradiation dose was 1.00 kJ / cm². 2 ~2.00kJ / cm 2 The irradiation dose was measured using a UVB radiometer, and the average UVB irradiation intensity was adjusted to 11.60 μW / cm² by adjusting the distance from the UV lamp. 2 According to the formula: UVB irradiation dose (mJ / cm²) 2 = Average light intensity (μW / cm) 2 Calculated by multiplying the irradiation time (s);

[0040] C. Incubation under constant light

[0041] Formulating with added Fe 2+ Mg 2+ Artemia dormant eggs were added to the saline solution containing GnRH-a at a concentration of 1.0 g / L to 3.0 g / L and incubated in a constant temperature incubator under constant light at a light intensity of 2000 Lx. The hatching rate was recorded after 1 to 3 days to determine the effect of removing diapause.

[0042] The brine preparation process is as follows: dilute aquaculture-specific sea salt (sodium chloride content >99%, brand not limited) with pure water to a concentration of 15‰-20‰; then add 5‰-15‰ Fe. 2+ and 5‰ to 15‰ Mg 2+ The final salt concentration was measured using a handheld ATC salinity refractometer to maintain the pH of the saline solution at 8.0–9.0. Finally, GnRH-a was added to the prepared saline solution to achieve a final concentration of 0.5–1.5 mg / L.

[0043] The incubation temperature for steps B and C is 26℃~30℃.

[0044] Example 2

[0045] (Amount of brine shrimp added, duration of initial light-protection treatment)

[0046] A. Collection and preservation of dormant brine shrimp eggs

[0047] Dormant eggs were collected from the brine shrimp breeding tank, washed with tap water, filtered, and dried. They were then stored in a -20℃ refrigerator for one week for low-temperature treatment. The stored brine shrimp dormant eggs were taken out of the -20℃ refrigerator, weighed and grouped according to Table 1, and a total of 27 portions were weighed and divided into nine experimental groups, with three replicates in each group.

[0048] B. Pretreatment of Artemia dormant eggs

[0049] The dormant Artemia eggs in each group from step A were placed in a smart light incubator, model GTOP-310B2, purchased from Zhejiang Top Instrument Co., Ltd., and subjected to the first dark treatment according to the time in Table 1 (1 for each group, 5 days of darkness, 7 days of darkness, and 9 days of darkness).

[0050] After the light-shielding treatment, the dormant Artemia eggs were transferred to an intelligent light incubator equipped with a Q-Lab / UVB-313EL ultraviolet lamp (purchased from Beijing Zhongyi Boteng Technology Co., Ltd.) for UVB irradiation treatment (parallel ultraviolet beam irradiation). The incubation temperature was set at 28℃. Using a UVB 297 ultraviolet radiometer (purchased from Xinbao Scientific Instruments Co., Ltd.), the average UVB irradiance was measured to be 11.60 μW / cm². 2The irradiation time was 1.5 days, during which the total irradiation dose was 1.50 J / cm². 2 .

[0051] The dormant eggs of the artichokes were then transferred back to the intelligent light incubator for a second dark treatment, which lasted for 2 days.

[0052] C. Incubation under constant light

[0053] Weigh out 18g of aquaculture-grade sea salt, 5.0g of ferrous sulfate, and 5.0g of magnesium sulfate, and add them to 1L of pure water to prepare a solution containing 5‰ Fe. 2+ and 5‰Mg 2+ The salinity of the brine was 18‰. The actual salinity was measured using a handheld ATC seawater salinity refractometer (specification: 0-28%) purchased from Guangzhou Huazhi Instrument Co., Ltd. The salinity was adjusted to 18‰ with pure water. At this point, the pH of the brine was kept between 8.0 and 9.0 (the pH can be adjusted with sterile HCl or NaOH solution). Finally, GnRH-a was added to the prepared brine to maintain the final concentration at 1.0 mg / L.

[0054] The Artemia eggs processed in step B were added according to the amounts in Table 1 and cultured in an incubator with the prepared saline solution for 2 days under constant light (light intensity 2000 Lx) at a temperature of 27℃. The hatching rate of Artemia eggs in each experimental group was calculated. The hatching rate was calculated using the dynamic microscopic examination method described in "Wang Jilin. Discussion on the Evaluation of Domestic Artemia Egg Hatching Rate Detection Methods [J]. Modern Fisheries Information". The hatching rate results are shown in Table 1 (the average value of each parallel group was taken and three significant figures were retained).

[0055] Table 1. Results of rapid dediapause hatching rate of Artemia salina eggs

[0056]

[0057] As can be seen from the results in Table 1, the optimal amount of Artemia eggs added is 2.0 g / L during the rapid dediapause and hatching process of Artemia dormant eggs; and the light-protection treatment combined with the need to shorten the overall hatching time is best done within 7 days.

[0058] Example 3

[0059] (UVB irradiation, GnRH-a dosage)

[0060] A. Collection and preservation of dormant brine shrimp eggs

[0061] Dormant eggs were collected from the brine shrimp breeding tank, washed with tap water, filtered, and dried. They were then stored in a -20℃ refrigerator for one week. The stored brine shrimp dormant eggs were taken out of the -20℃ refrigerator and accurately weighed 2.0g each. A total of 27 portions were weighed and grouped according to Table 2, forming nine experimental groups with three replicates in each group.

[0062] B. Pretreatment of Artemia dormant eggs

[0063] The dormant Artemia eggs from each group in step A were placed in a smart light incubator (model GTOP-310B2) purchased from Zhejiang Top Instrument Co., Ltd., and subjected to initial light-shielding treatment for 7 days.

[0064] After the light-shielding treatment, the dormant Artemia eggs were transferred to an intelligent light incubator equipped with Q-Lab / UVB-313EL ultraviolet lamps purchased from Beijing Zhongyi Boteng Technology Co., Ltd., for UVB irradiation treatment (parallel ultraviolet beam irradiation). The incubation temperature was set at 28℃. Group A was irradiated for 1 day, Group B for 1.5 days, and Group C for 2 days. At this time, the total irradiation dose for Groups A, B, and C was 1.00 J / cm³. 2 1.50 J / cm 2 2.00 J / cm 2 .

[0065] The dormant eggs of the artichokes were then transferred back to the intelligent light incubator for a second dark treatment, which lasted for 2 days.

[0066] C. Incubation under constant light

[0067] Weigh out 20g of aquaculture-grade sea salt, 5.0g of ferrous sulfate, and 5.0g of magnesium sulfate, and add them to 1L of pure water to prepare a solution containing 5‰ Fe. 2+ and 5‰Mg 2+ The concentration of the saline solution was 20‰, and the pH of the saline solution was 8.6. Finally, GnRH-a was added to the prepared saline solution so that the final concentration of each group met the requirements of Table 2.

[0068] The artichoke eggs processed in step B were added to the saline solution prepared according to Table 2 at a concentration of 2.0 g / L and cultured in an incubator under constant light (light intensity 2000 Lx) for 2 days at a temperature of 27℃. The hatching rate of the artichoke eggs in each experimental group was calculated, and the results are shown in Table 2.

[0069] Table 2. Results of rapid dediapause hatching rate of Artemia salina eggs

[0070]

[0071] As shown in Table 2, during the rapid dediapause and hatching process of Artemia salina eggs, the Artemia salina eggs were exposed to a UVB irradiation dose of 1.50 J / cm². 2 The effect was better when the final concentration of GnRH-a was 1.0 g / L. The hatching rate of Artemia eggs decreased rapidly when the final concentration of GnRH-a was 1.5 mg / L, which may be because high concentration of GnRH-a can delay or even inhibit its dediapause.

[0072] Example 4

[0073] (Cultivation temperature and incubation time)

[0074] A. Collection and preservation of dormant brine shrimp eggs

[0075] Dormant eggs were collected from the brine shrimp breeding tank, washed with tap water, filtered, and dried. They were then stored in a -20℃ refrigerator for one week. The stored brine shrimp dormant eggs were taken out of the -20℃ refrigerator and accurately weighed 2.0g each. A total of 27 portions were weighed and grouped according to Table 2, forming nine experimental groups with three replicates in each group.

[0076] B. Pretreatment of Artemia dormant eggs

[0077] The dormant Artemia eggs from each group in step A were placed in a smart light incubator (model GTOP-310B2) purchased from Zhejiang Top Instrument Co., Ltd., and subjected to an initial dark treatment for 7 days.

[0078] After the light-shielding treatment, the dormant Artemia eggs were transferred to an intelligent light incubator equipped with a Q-Lab / UVB-313EL ultraviolet lamp purchased from Beijing Zhongyi Boteng Technology Co., Ltd., and subjected to UVB irradiation treatment (parallel ultraviolet beam irradiation) for 1.5 days. The incubation temperature was set according to Table 3, and the total irradiation dose was 1.50 J / cm². 2 .

[0079] The dormant eggs of Artemia were then transferred back to the intelligent light incubator for a second dark treatment for 2 days.

[0080] C. Incubation under constant light

[0081] Weigh out 20g of aquaculture-grade sea salt, 10.0g of ferrous sulfate, and 10.0g of magnesium sulfate, and add them to 1L of pure water to prepare a solution containing 10‰ Fe. 2+ and 10‰ Mg 2+ The concentration of the brine was 20‰, and the pH of the brine was 8.8. Finally, GnRH-a solution was added to the prepared brine to maintain its concentration at 1.0 mg / L.

[0082] The artichoke eggs processed in step B were added to the prepared saline solution at a concentration of 2.0 g / L and cultured in an incubator under constant light (light intensity 2000 Lx) according to the method in Table 3. The hatching rate of artichoke eggs in each experimental group was calculated, and the hatching rate results are shown in Table 3.

[0083] Table 3. Results of rapid dediapause hatching rate of Artemia salina eggs

[0084]

[0085]

[0086] As can be seen from the results in Table 3, the optimal culture temperature for Artemia eggs during the rapid dediapause and hatching process is 28℃. After being protected from light and irradiated with UVB, the hatching rate fluctuates little after 2-3 days of culture under normal light. The preferred culture time is more than 2 days.

[0087] Example 5

[0088] (Fe 2+ Mg 2+ (Comparison of content and the effect of different sources of artemisia on diapause)

[0089] A. Collection and preservation of dormant brine shrimp eggs

[0090] Dormant eggs were collected from the brine shrimp breeding tank, washed with tap water, filtered, and dried. They were then stored in a -20℃ refrigerator for one week. The stored brine shrimp dormant eggs were taken out of the -20℃ refrigerator and accurately weighed 2.0g each. A total of 27 portions were weighed and grouped according to Table 2, forming nine experimental groups with three replicates in each group.

[0091] B. Pretreatment of Artemia dormant eggs

[0092] The dormant Artemia eggs from each group in step A were placed in a smart light incubator (model GTOP-310B2) purchased from Zhejiang Top Instrument Co., Ltd., and subjected to an initial dark treatment for 7 days.

[0093] After the light-protection treatment, the dormant Artemia eggs were transferred to an intelligent light incubator equipped with a Q-Lab / UVB-313EL ultraviolet germicidal lamp purchased from Beijing Zhongyi Boteng Technology Co., Ltd., and treated with UVB irradiation (parallel ultraviolet beam irradiation) for 1.5 days at a culture temperature of 27℃. The total irradiation dose was 1.50 J / cm². 2 .

[0094] The dormant eggs of Artemia were transferred back to the intelligent light incubator for a second dark treatment, and then the dormant eggs after irradiation were kept in the dark for 2 days.

[0095] C. Incubation under constant light

[0096] Weigh 20g of aquaculture-grade sea salt and add it to 1L of pure water to prepare a 20‰ salt solution. According to Table 4, Fe... 2+ Mg 2+ Ferrous sulfate and magnesium sulfate were added to the solution, bringing the pH of the brine to between 8.5 and 9.0. Finally, GnRH-a solution was added to the prepared brine to maintain its concentration at 1.0 g / L.

[0097] The Artemia eggs processed in step B were added to the above-prepared saline solution at a dosage of 2.0 g / L and cultured in an incubator under constant light (light intensity 2000 Lx) for 2 days. The hatching rate of Artemia eggs in each experimental group was calculated, and the hatching rate results are shown in Table 4.

[0098] Table 4. Results of rapid dediapause hatching rates of dormant eggs from different sources of Artemia salina.

[0099]

[0100]

[0101] As can be seen from the results in Table 4, the preferred conditions of this invention differ only slightly for dormant Artemia eggs from different sources, while Fe 2+ Mg 2+ The effect of Fe content on hatching rate shows a trend of "first increasing and then decreasing". Overall, the Fe content in the brine used for hatching... 2+ Mg 2+ When the concentration is 10‰, the hatching rate is relatively high.

[0102] By reviewing relevant literature and patents, it was found that existing technologies for removing diapause from Artemia eggs have varying degrees of drawbacks. Firstly, the diapause removal method reported in "Yang Na, Bian Bozhong, Li Mingren. Study on Hatching Characteristics of Artemia Eggs in China [J]. Journal of Fisheries of China" is time-consuming, generally requiring 2-3 months. Furthermore, the commonly used diapause removal reagents, 3% H2O2 and 3% formaldehyde solution, have no effect on ending the diapause state of dormant eggs produced by indoor-cultured Artemia, but are effective for wild-collected Artemia eggs, thus limiting their applicability. Secondly, the method for improving the hatching rate disclosed in CN 106577408A, entitled "A Preparation of Decapitated Artemia Eggs for Improving Hatching Rate," uses a biological enzymatic hydrolysis method for decapitation, requiring numerous reagents and complex steps, including multiple enzyme preparations, and also incorporating microbial fermentation experiments, significantly increasing experimental costs. The invention disclosed in CN104273098A, entitled "A Method for Hatching Diapause Artemia Eggs under Ultraviolet Light," also requires the eggs to be shed. However, it differs in that the shedding agent used is a chemical reagent such as hypochlorous acid, which can easily damage the eggs. The eggs used are commercially available, and the same effect cannot be achieved with dormant eggs produced by indoor-cultured artemia. Our research team used the method described in CN104273098A to experiment with dormant eggs produced by parthenogenetic artemia in our laboratory, and found that the hatching rate was only about 75%.

[0103] Using the method of this invention, the diapause time of dormant brine shrimp eggs cultured indoors is shortened to less than 20 days, and the hatching rate is increased to 92.5%.

[0104] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. The same or similar parts between the various embodiments can be referred to each other.

[0105] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for rapidly breaking diapause in dormant brine shrimp eggs cultured indoors and improving hatching rate, characterized in that, Includes the following steps: A. Collection and preservation of dormant brine shrimp eggs The collected Artemia dormant eggs were washed, filtered, and dried, and then stored at -20°C for one week for low-temperature treatment; the Artemia dormant eggs were dormant eggs produced by laboratory-cultured Artemia. B. Pretreatment of Artemia dormant eggs First, the dormant eggs of Artemia after low-temperature treatment were placed in an incubator for the first light-proof and dark treatment for 7 days; then, the dormant eggs of Artemia were transferred to an incubator equipped with a UVB lamp for UVB irradiation treatment for 1-2 days; then the dormant eggs of Artemia were transferred back to the incubator for a second light-proof and dark treatment for 2 days. The incubator is a GTOP-310B2, BZ-GTOP-300Y, or NB-KRG-400A type intelligent light incubator; the UVB irradiation treatment uses a parallel ultraviolet beam, and the UVB irradiation equipment used is a GPH873T5L / 40W or Q-Lab / UVB-313EL type ultraviolet lamp; the UVB irradiation dose is 1.00 kJ / cm². 2 ~2.00 kJ / cm 2 The irradiation dose was measured using a UVB radiometer, and the average UVB irradiation intensity was adjusted to 11.60 μW / cm² by adjusting the distance from the UV lamp. 2 According to the formula: UVB irradiation dose (mJ / cm²) 2 = Average light intensity (μW / cm) 2 Calculated by multiplying the irradiation time (s); C. Light-based cultivation and incubation Formulating with added Fe 2+ Mg 2+ Artemia dormant eggs were added to the saline solution containing GnRH-a to achieve a concentration of 1.0 g / L to 3.0 g / L. The eggs were then incubated in a constant temperature incubator under light at an intensity of 2000 Lx. The hatching rate was recorded after 1 to 2 days to determine the effectiveness of diapause removal. The brine preparation process is as follows: dilute aquaculture-specific sea salt with pure water to a concentration of 15‰ to 20‰ by mass-volume ratio; then add 5‰ to 15‰ Fe. 2+ and 5‰ to 15‰ Mg 2+ The final salt concentration was measured using a handheld ATC salinity refractometer to maintain the pH of the brine at 8.0–9.0; then GnRH-a was added to bring the final concentration to 1.5 mg / L. The incubation temperature for both steps B and C is 26℃~30℃; The dormant eggs of Artemia are parthenogenetic Artemia eggs.

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