Cultivation method of pterocladia caerulea

Abstract

The invention discloses a method for cultivating pterocladia caerulea. Relates to the field of seaweed industrialized seedling production. The cultivation method of the pterocladia capillipes comprises the following steps: S1, taking the pterocladia capillipes, and performing disinfection and crushing treatment to obtain pterocladia capillipes frond particles; s2, spreading the Prassica rapa frond particles on an adhesion substrate; s3, placing the substratum in an oligotrophic water body for standing culture until stolons and rhizoids grow on the Prassica rapa frond particles and are firmly attached to the substratum; and S4, transferring the substratum attached with the brassica rapa frond into a seawater environment for culturing. According to the method disclosed by the invention, by crushing into particles and optimizing the culture conditions, the seedling raising period from sowing to complete attachment of the pterocladia caerulea is shortened to about 40 days and is shortened by more than 33% compared with that of a traditional tissue culture seedling raising method (about 60 days), the bottlenecks of long seedling raising period and low efficiency in the traditional technology are solved, and the time requirement of factory-like batch seedling raising can be met.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of kelp industrial breeding, and in particular to a Pterocladiella capillacea cultivation method. BACKGROUND

[0002] Pterocladiella capillacea, as a representative species of the Pterocladiellaceae family in the order of Gelidiales in the Rhodophyta phylum, occupies a unique position in the marine ecosystem due to its biological characteristics and ecological adaptability. This species typically exhibits a slender branching structure, with algal bodies ranging in color from purple-red to light brown, and a height of several centimeters to over ten centimeters. The dense branching and high elasticity of Pterocladiella capillacea enable it to remain stable and attached even under the impact of waves in the intertidal zone. As a typical representative of subtropical seaweed, Pterocladiella capillacea prefers shallow sea areas with water temperatures between 15–25℃ and has a certain tolerance to changes in salinity, which allows it to form dominant populations in the intertidal reef areas of Fujian, Guangdong, and Guangxi in China. Its life cycle includes multiple stages such as sporophytes, gametophytes, and carposporophytes, and the demands for light, nutrient salt concentration, and water flow velocity differ between stages, which poses natural challenges to artificial cultivation.

[0003] Agar, as the main polysaccharide component of the cell wall of Pterocladiella capillacea, has unique gel properties that make it irreplaceable in the fields of biomedical, food industry, and cosmetics. In the biomedical field, agar is often used as a cell culture scaffold, a drug release carrier, and a matrix for tissue engineering materials due to its good biocompatibility and biodegradability. In the food industry, it is widely used in the production of products such as jelly, yogurt, and meat products as a natural thickening agent, stabilizer, and coagulant. In the cosmetics industry, the moisturizing properties and colloidal stability of agar make it an important ingredient in high-end skincare products. Compared with other seaweed-derived gums, the advantages of Pterocladiella capillacea agar not only lie in its gel strength and temperature stability, but also in its low ash content and high transparency, which significantly improve the quality of end products. This makes Pterocladiella capillacea a preferred raw material at various stages of the industrial chain.

[0004] The current development of agar industry is facing a profound contradiction between resource supply and market demand. With the rapid expansion of the market for biomedical materials and the rise of the trend of healthy food consumption, the global demand for agar is growing at an average annual rate of two digits, while the supply of raw materials is subject to the limitation of wild seaweed resources. As a major producer and consumer of agar, China's industry expansion is directly dependent on the harvesting of wild Gracilaria lemaneiformis in the southeast coast. This extensive way of resource acquisition has caused a series of ecological and economic problems. At the ecological level, over-harvesting has led to the destruction of intertidal algal community structure, which in turn affects the diversity of small marine organisms that use seaweed as habitat and weakens the ecological protection function of the coastal zone. At the economic level, the sharp decline in wild resources has pushed up the cost of raw material procurement. Some processing enterprises have been forced to reduce production capacity or even exit the market due to their inability to bear the high cost of raw materials, forming a vicious cycle of "resource reduction - price increase - intensified exploitation". This situation not only restricts the sustainable development of the agar industry, but also poses a potential threat to China's strategic position in the global competition for seaweed resources.

[0005] The breakthrough of artificial cultivation technology is considered as the key path to break this dilemma, and the seedling stage as the starting point of the cultivation industry chain, its technical maturity directly determines the feasibility of industrialization. At present, the seedling technology of Gracilaria lemaneiformis mainly includes spore seedling and tissue culture seedling, both of which have insurmountable technical bottlenecks. Spore seedling technology is based on the natural reproduction law, through inducing algae to release spores and adhere to germinate on artificial substrate, finally cultivating into seedlings. However, the release of spores of Gracilaria lemaneiformis is regulated by multiple environmental factors such as light cycle, temperature fluctuation and nutrient salt level. Although spore release can be achieved in laboratory conditions, it is difficult to stably control the release efficiency in large-scale production, resulting in significant differences between batches of seedlings. At the same time, the period from spore germination to seedling is as long as 3-4 months, during which high-precision water quality parameters need to be maintained, resulting in high management cost. Tissue culture seedling technology achieves rapid propagation through in vitro tissue culture, shortening the seedling period to about 2 months, but the core problem lies in the prevention and control of contamination by impurities. In open cultivation systems, diatoms and green algae compete with Gracilaria lemaneiformis seedlings for survival resources. Once pollution breaks out, the whole batch of seedlings may be wiped out. This high-risk characteristic makes enterprises cautious about large-scale application.

[0006] Therefore, it is urgent to develop a factory artificial cultivation method of Gracilaria lemaneiformis with rapid propagation, open cultivation and low pollution risk to solve the problems existing in the current artificial cultivation. SUMMARY

[0007] The purpose of the present application is to develop a factory artificial cultivation method of Gracilaria lemaneiformis with rapid propagation, open cultivation and low pollution risk.

[0008] The first aspect of the present application is:

[0009] Provided is a breeding method of Sargassum siliquastrum.

[0010] A second aspect of the present application is:

[0011] The breeding method of Sargassum siliquastrum.

[0012] Specifically, the technical solution adopted according to the first aspect of the present application is:

[0013] A breeding method of Sargassum siliquastrum, comprising the following steps:

[0014] S1: Sargassum siliquastrum is taken, disinfected and crushed to obtain Sargassum siliquastrum thallus particles;

[0015] S2: The Sargassum siliquastrum thallus particles are sown on an attachment base;

[0016] S3: The attachment base is placed in an oligotrophic water body for static culture until the Sargassum siliquastrum thallus particles grow outstretched stems and rhizoids and firmly attach to the attachment base;

[0017] S4: The attachment base with the Sargassum siliquastrum thallus attached is transferred to a seawater environment for cultivation;

[0018] The oligotrophic water body is prepared by the following method: Zhang's Gracilaria is inoculated in an indoor cement pool at an inoculation density of 0.8-1.2 kg / m 3 , and the oligotrophic water body is obtained after 12-18 days of culture.

[0019] According to an embodiment of the present application, one of the technical solutions has at least one of the following advantages or beneficial effects:

[0020] The present application shortens the cultivation period of Sargassum siliquastrum from sowing to complete attachment to about 40 days, which is more than 33% shorter than the traditional tissue culture method (about 60 days), solves the bottleneck of long cultivation period and low efficiency in the traditional technology, and meets the time requirements of factory-scale seedling cultivation.

[0021] Among them, the oligotrophic environment (the concentration of nitrogen, phosphorus and other nutrient salts is significantly lower than that of ordinary seawater) constructed by Zhang's Gracilaria pre-absorbing water body nutrients can specifically inhibit the growth of miscellaneous algae, while the Sargassum siliquastrum particles can still develop normally due to their low nutrient salt tolerance. In actual cultivation, the biomass of miscellaneous algae is much lower than that in the traditional eutrophic environment, thereby improving the purity of Sargassum siliquastrum seedlings and solving the core problem of seedling pollution caused by competition with miscellaneous algae in large-scale cultivation.

[0022] In the present application, the broken-up Gracilaria Lemanei is matched with the gap of the attachment base, and can grow out stolon and rhizoid quickly during the static culture, so as to be firmly attached to the attachment base, and the effect of traditional spore seedling is significantly improved, and finally the high survival rate of seedling from indoor cultivation to outdoor cultivation is ensured. After the indoor cultivation is completed, the attachment base with the Gracilaria Lemanei algae attached thereto can be transferred to any seawater environment for cultivation.

[0023] According to an embodiment of the present application, the disinfectant for disinfection is trichloroisocyanuric acid, preferably, the concentration of the trichloroisocyanuric acid is 0.8-1.2 g / L, and the time for disinfection is 0.5-1.5 min.

[0024] According to an embodiment of the present application, the disinfection is to soak the Gracilaria Lemanei in the disinfectant for disinfection, preferably, the soaking time for disinfection is 0.5-1.5 min. The present application limits the high-efficiency and low-toxicity disinfection parameters, avoids the damage to the algae while killing the attached microorganisms, and 0.8-1.2 g / L trichloroisocyanuric acid soaking for 0.5-1.5 min can effectively disinfect, and the short-time treatment reduces the influence on the activity of the algae.

[0025] According to an embodiment of the present application, the disinfection further comprises the following steps: soaking and stirring the Gracilaria Lemanei after disinfection with seawater, and repeating the seawater soaking for 4-5 times. Preferably, the seawater soaking time is 20-40 s for the first 2 times, and 1.5-3 min for the last 2-3 times. The present application controls the seawater soaking time in stages to completely remove the residual disinfectant and avoid the continuous toxicity to the algae.

[0026] According to an embodiment of the present application, the device for breaking up is a crusher.

[0027] According to an embodiment of the present application, the specific operation of the breaking-up treatment is as follows: the Gracilaria Lemanei is placed in the crusher, seawater is added to the crusher just to cover the Gracilaria Lemanei, the crusher is stopped for 0.5-0.6 min every 1-1.2 min of operation, and the cycle is repeated for 2-4 times before the filtration treatment. The present application controls the temperature rise of the device during the breaking-up process to avoid the high temperature from damaging the activity of the algae.

[0028] According to an embodiment of the present application, the filtration treatment in the breaking-up treatment is to use a filter screen with a pore size of 0.3-0.7 cm. The present application limits the pore size of the filter screen to limit the size of the algae particles, so as to adapt to the gap of the attachment base and improve the attachment efficiency.

[0029] According to an embodiment of the present application, the pore size of the filter screen is 0.5 cm. The particles obtained by using the filter screen with a pore size of 0.5 cm can be clamped in the gap between the seedling curtain ropes, so as to avoid falling off.

[0030] According to one embodiment of the present invention, the attachment substrate is a fiber-woven seedling curtain. The present invention uses a seawater-resistant blended material to ensure the stability and reusability of the attachment substrate.

[0031] According to one embodiment of the present invention, the fiber includes at least one of vinylon and polyethylene.

[0032] According to one embodiment of the present invention, the size of the seedling curtain is 20-25cm × 50-55cm. A suitable size for the seedling curtain allows it to be adapted to factory-produced cement ponds.

[0033] According to one embodiment of the present invention, in step S2, the distribution density of the *Pseudo-Gynostemma pentaphyllum* thallus particles on the substrate is 100-300 particles / dm². Overlapping of the *Pseudo-Gynostemma pentaphyllum* thallus particles should be avoided, otherwise it will affect the adhesion to the substrate.

[0034] According to one embodiment of the present invention, the temperature for static cultivation is 20-30°C; and / or the light intensity is 2500-3300 Lux. Insufficient light is detrimental to algal development, while excessive light easily leads to the growth of unwanted algae. Appropriate light intensity is beneficial to algal development and inhibits unwanted algae.

[0035] According to one embodiment of the present invention, the temperature for static culture is preferably 25-26°C, and the light intensity is preferably 2500-2800 Lux.

[0036] According to one embodiment of the present invention, the static culture time is 35-40 days.

[0037] According to one embodiment of the present invention, the water level fluctuation during the static culture period does not exceed ±2 cm. The water level fluctuation refers to the range of water level changes in the aquaculture environment.

[0038] According to one embodiment of the present invention, the substrate is fixed in the indoor cement tank by a bracket, the bracket being 5-10 cm below the water surface. Preferably, the bracket is constructed of PVC pipe. The bracket's height being 5-10 cm below the water surface ensures the substrate is completely submerged, and the appropriate water depth places the algal particles in the upper layer of the water, allowing for more sufficient light and dissolved oxygen.

[0039] According to one embodiment of the present invention, the timing for transferring the thallus to a seawater environment for cultivation is when any *Pterocarya pseudobulb* thallus particle grows three or more stolons with a length of 0.8-1.2 cm.

[0040] Specifically, the technical solution adopted according to the second aspect of the present invention is as follows:

[0041] An attached base for cultivating Gracilaria chouiae, which is knitted by a mixture of vinylon and polyethylene, and has a size of 20-25 cm x 50-55 cm.

[0042] Another aspect of the present application also relates to an industrialized seedling system applying the cultivating method of Gracilaria chouiae, which comprises an indoor cement pool, a Zhang's Gracilaria inoculation unit, an attached base support and an environmental control unit, the Zhang's Gracilaria inoculation unit is used for inoculating Zhang's Gracilaria in the indoor cement pool, and the environmental control unit is used for regulating temperature and light.

[0043] Additional aspects and advantages of the present application will be in part apparent and in part pointed out hereinafter. BRIEF DESCRIPTION OF DRAWINGS

[0044] The above and / or additional aspects and advantages of the present application will become apparent and be readily appreciated from the following description, including the appended drawings.

[0045] Figure 1 A photograph of the seedling screen to which Gracilaria chouiae granules were scattered in Example 1.

[0046] Figure 2 A photograph of Gracilaria chouiae cultivated in an oligotrophic environment in Example 1.

[0047] Figure 3 A photograph of Gracilaria chouiae cultivated in a eutrophic environment in the comparative example. DETAILED DESCRIPTION

[0048] The words "preferred" and "preferably" in this document refer to embodiments of the application that can provide certain benefits under certain circumstances. However, other embodiments can also be preferred, or preferred in other circumstances. Additionally, the recitation of one or more preferred embodiments does not imply that other embodiments are not useful, and is not intended to exclude those other embodiments from the scope of the application.

[0049] When a numerical range is disclosed herein, the range is to be construed as continuous, and to include each and every value and sub-range within the range. Further, when a range is disclosed, the range is inclusive of the minimum and maximum values, and of each integer within the range. Further, when a plurality of ranges is provided, the ranges can be combined. In other words, unless otherwise indicated, all ranges disclosed herein are to be understood as encompassing any and all sub-ranges subsumed therein.

[0050] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. 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 of ordinary skill in the art without creative effort are within the scope of the present invention.

[0051] Unless otherwise specified, the reagents, methods and equipment used in this invention are all conventional reagents, methods and equipment in this technical field.

[0052] Example 1

[0053] A method for cultivating *Gnaphalium affine* includes the following steps:

[0054] S1. Rinse the *Syngonium spp.* clean. Immerse the cleaned *Syngonium spp.* in a 1 g / L solution of trichloroisocyanuric acid in seawater for 1 minute to disinfect it. After disinfection, remove the *Syngonium spp.* and soak and stir it in seawater. Repeat this process 4 times with fresh seawater to remove any residual disinfectant from the algae. The first and second rinses should not exceed 30 seconds, and the third and fourth rinses should be 2 minutes each. Then, place the disinfected *Syngonium spp.* into a pulverizer and add seawater just enough to cover the algae. Stop the pulverizer for 0.5 minutes every 1 minute of operation to avoid overheating and damaging the algae. After three cycles of operation, filter the algae through a 0.5 cm pore size filter to obtain *Syngonium spp.* algae particles.

[0055] S2. The *Pterocarya pseudobulb* thallus particles are evenly spread on the seedling tray (a 20cm x 50cm seedling board woven from vinylon-polyethylene blended rope) as evenly as possible, ensuring the thallus particles do not overlap. A picture of the seedling tray with the *Pterocarya pseudobulb* thallus particles is shown below. Figure 1 As shown; Gracilaria zhangi cultured in seawater was added to an indoor cement tank, with a density of 1 kg / m³. 3 After about 15 days of cultivation, an oligotrophic water body is obtained; a rectangular support frame is built using PVC pipes with a diameter of 3.2cm, which is 8cm lower than the water surface of the cement pond. The seedling curtain is tied to the support frame with cable ties to fix the seedling curtain.

[0056] S3 seedling curtains inoculated with *Pseudobulbus simulans* thallus particles were placed in oligotrophic water and cultured statically for 40 days. Indoor culture conditions were 25℃ and 2800 Lux light, until most of the *Pseudobulbus simulans* thallus particles had grown multiple stolons about 1 cm long, which firmly attached themselves to the seedling ropes of the seedling curtain via rhizoids, resulting in... Figure 2 The image shows *Gnaphalium affine* cultivated in an oligotrophic environment.

[0057] S4 transferred seedlings with *Gnaphalium affine* attached to the curtains to a marine environment for cultivation.

[0058] From Figure 2 It can be seen that under the oligotrophic environment, only the firmly attached Gracilaria chouae is on the seedling screen, and there is no other miscellaneous algae, indicating that the oligotrophic environment obtained under specific culture conditions can specifically inhibit the growth of miscellaneous algae, and the Gracilaria chouae granules can still develop normally due to the low-nutrient salt tolerance characteristics, thereby improving the purity of the Gracilaria chouae seedlings and solving the core problem of "miscellaneous algae competition leading to seedling contamination" in large-scale seedling cultivation in factories.

[0059] Example 2

[0060] A Gracilaria chouae cultivation method, comprising the following steps:

[0061] S1 Clean the Gracilaria chouae, immerse the cleaned Gracilaria chouae in a 1.2 g / L seawater solution of trichloroisocyanuric acid for 0.5 min for disinfection, then take out the disinfected Gracilaria chouae, soak and stir in seawater, take out again, replace with new seawater, and repeat for 4 times to wash off the disinfectant residues on the algae, wherein the cleaning time of the first and second times is not more than 30 s, and the cleaning time of the third and fourth times is 1.5 min; then, put the disinfected Gracilaria chouae into a pulverizer and add seawater just covering the Gracilaria chouae, stop the pulverizer every 1 min for 0.5 min to avoid high temperature damaging the algae, and filter with a filter screen with a pore size of 0.3 cm after three starts to obtain Gracilaria chouae algae granules;

[0062] S2 Spread the Gracilaria chouae algae granules as evenly as possible on the seedling screen (the seedling screen is a 20 cm*50 cm seedling plate woven by Vinyon-polyethylene blended rope), and try not to overlap between the algae granules; add Zhang's Gracilaria in seawater in a cement pool in the room, the density of Zhang's Gracilaria is 0.8 kg / m 3 around, and cultivate for 18 days to obtain an oligotrophic water body; use a PVC pipe with a diameter of 3.2 cm to build a rectangular cuboid support 6 cm lower than the water surface of the cement pool, and bind the seedling screen to the support to fix the seedling screen;

[0063] S3 Place the seedling screen with Gracilaria chouae algae granules in the oligotrophic water body and incubate for 40 days, the indoor culture conditions are temperature of 25℃ and illumination of 2800Lux, until most of the Gracilaria chouae algae granules grow out of a plurality of 1 cm long creeping stems, and firmly attach to the seedling ropes of the seedling screen through rhizoids;

[0064] S4 Transfer the seedling screen with Gracilaria chouae to seawater environment for cultivation.

[0065] Comparative example

[0066] The difference between the comparative example and example 1 is that in the comparative example, the Gracilaria chouae is cultivated in eutrophic seawater.

[0067] Specifically:

[0068] A method for cultivating a Gracilaria dura, comprising the following steps:

[0069] S1: Clean Gracilaria dura, immerse the cleaned Gracilaria dura in a seawater solution of 1 g / L trichloroisocyanuric acid for 1 min for disinfection, take out the disinfected Gracilaria dura, immerse and stir in seawater, take out again, replace with new seawater, and repeat for 4 times to wash away the disinfectant remaining on the algal body, wherein the cleaning time of the first and second times is not more than 30 s, and the cleaning time of the third and fourth times is 2 min; then, put the disinfected Gracilaria dura into a pulverizer and add seawater just covering the Gracilaria dura, stop the pulverizer every 1 min for 0.5 min to avoid damage to the algal body due to high temperature, and filter with a filter screen with a pore size of 0.5 cm after starting the pulverizer for three times to obtain Gracilaria dura algal body particles;

[0070] S2: Spread the Gracilaria dura algal body particles as evenly as possible on a seedling screen (the seedling screen is a 20 cm*50 cm seedling plate woven by a Vinyon-polyethylene blended rope), and try not to overlap the algal body particles; add seawater in a cement pool in a room, and stand for 15 days to obtain a nutrient-rich water body; use a PVC pipe with a diameter of 3.2 cm to build a cuboid support 8 cm lower than the water surface of the cement pool, and bind the seedling screen on the support to fix the seedling screen;

[0071] S3: Place the seedling screen with the Gracilaria dura algal body particles spread thereon in the nutrient-rich water body and stand for culture for 40 days, the indoor culture conditions are a temperature of 25℃ and illumination of 2800 Lux, and Gracilaria dura cultured in a nutrient-rich environment as shown in Figure 3 is obtained;

[0072] S4: Transfer the seedling screen with the Gracilaria dura attached thereto to a seawater environment for cultivation.

[0073] It can be seen from Figure 3 that in addition to the Gracilaria dura, there are a large amount of miscellaneous seaweed in the seedling screen in the nutrient-rich environment, and the seedling screen is seriously contaminated, which is not conducive to batch seedling culture.

[0074] The above are only embodiments of the present application, and do not limit the patent scope of the present application, and any equivalent transformation or direct or indirect application in related technical fields using the content of the present application is also included in the patent protection scope of the present application.

Claims

1. A method for breeding a plant of the species Brassica rapa, characterized by: The method comprises the following steps: S1: obtaining Gracilaria chouae algae particles by disinfecting and crushing Gracilaria chouae; S2: spreading the Gracilaria chouae algae particles on an attachment base; S3: placing the attachment base in an oligotrophic water body for static culture until the Gracilaria chouae algae particles grow out of stolons and rhizoids and firmly attach to the attachment base; S4: transferring the attachment base with the Gracilaria chouae algae to a seawater environment for cultivation. The oligotrophic water body is prepared by the following method: inoculating Zhangjiang Gracilaria in a cement pool in a room, the inoculation density being 0.8-1.2 kg / m 3 , and obtaining the oligotrophic water body after 12-18 days of culture.

2. The breeding method of the ikejikoucai according to claim 1, characterized in that: The disinfectant used for disinfection is trichloroisocyanuric acid, preferably, the concentration of the trichloroisocyanuric acid is 0.8-1.2 g / L, and the time for disinfection is 0.5-1.5 min.

3. The breeding method of the ikejikoucai according to claim 1, characterized in that: The specific operation of the crushing treatment is as follows: placing Gracilaria chouae in a crusher, adding seawater just covering the Gracilaria chouae in the crusher, stopping the crusher every 1-1.2 min for 0.5-0.6 min, and performing filtration treatment after 2-4 cycles of circulation.

4. The breeding method of the ikejikoucai according to claim 3, characterized in that: The filtration treatment in the crushing treatment is filtration using a filter screen with a pore size of 0.3-0.7 cm.

5. The breeding method of the ikejikoucai according to claim 1, characterized in that: The attachment base is a fiber-woven nursery curtain.

6. The breeding method of the ikejikoucai according to claim 1, characterized in that: The temperature of the static culture is 20-30℃; and / or, the light intensity is 2500-3300 Lux.

7. The breeding method of the ikejikoucai according to claim 1, characterized in that: The time of the static culture is 35-40 days.

8. The breeding method of the ikejikoucai according to claim 1, characterized in that: The attachment base is fixed in the indoor cement pool by a support, and the height of the support is 5-10 cm lower than the water surface.

9. The breeding method of the ike-nagaimo according to claim 1, characterized by: The timing of the transfer to the seawater environment for cultivation is when any Gracilaria chouae algae particle grows out of 3 or more stolons with a length of 0.8-1.2 cm.

10. An industrialized seedling raising system for applying the Brassica napus L. breeding method according to claims 1-9, characterized in that: It comprises an indoor cement pool, a Zhang's Gracilaria inoculation unit, an attachment base support, and an environmental control unit, the Zhang's Gracilaria inoculation unit is used for inoculating Zhang's Gracilaria in the indoor cement pool, and the environmental control unit is used for regulating temperature and light.

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