50 results about "Crassostrea hongkongensis" patented technology

The invention discloses a preparation method of a Crassostrea hongkongensis fixation medium. The preparation method comprises the following steps that (1) a plurality of rectangular plastic sheets are made and a round hole with the radius being 3-5 mm is formed in each plastic sheet, wherein the length of the plastic sheets is 10-20 cm, the width of the plastic sheets is 2-5 cm and the thickness of the plastic sheets is 1-3 cm; (2) the plastic sheets are strung through a rope, a plastic tube is arranged between every two adjacent plastic sheets, the plastic tubes are arranged on the rope in a sleeved mode, and then the preparation process of the Crassostrea hongkongensis fixation medium is completed. The invention further discloses the Crassostrea hongkongensis fixation medium and an application method of the Crassostrea hongkongensis fixation medium. After being processed through oxalic acid, fresh water and the like, the Crassostrea hongkongensis fixation medium is used as an eyespot larva collection fixation medium; the Crassostrea hongkongensis fixation medium increases the metamorphosis rate of Crassostrea hongkongensis larvas, accelerates juvenile mollusk growth, improves the juvenile mollusk survival ability and increases the juvenile mollusk yield, has the advantages of being capable of being recycled multiple times, light, convenient to use, good in corrosion resistance and the like and is worthy of popularization and application.

The invention discloses a method suitable for artificial breeding of CrassOstrea hongkongensis in a sea area in the north. The method suitable for artificial breeding of CrassOstrea hongkongensis in the sea area in the north is characterized by comprising the steps that firstly, CrassOstrea hongkongensis adult shellfish with a shell higher than 10 cm are selected, the CrassOstrea hongkongensis adult shellfish are placed in breeding ponds in high-temperature summer (July and August) with the density of 40-50 pieces / m<3> under the conditions that the water temperature ranges from 26 DEG C to 30 DEG C and the salinity ranges from 25 to 32, artificial oviposition and spermiation are conducted after the CrassOstrea hongkongensis adult shellfish are bred for 10 days to 15 days, spawn and sperms are placed in a breeding pond containing filtered sea water to be discharged naturally after being dried in the shade for five hours to eight hours, or the spawn and the sperms are obtained after the male shellfish and female shellfish are distinguished manually, and then artificial insemination is conducted; germ cells are hatched for 18 hours to 20 hours under the condition that the water temperature ranges from 26 DEG C to 32 DEG C so that D type larvas can be obtained, the D type larvas are placed in ponds to be bred with the density of 6-8 pieces / m<3>, when 50%-80% of the D type larvas become eyespot larvas after the D type larvas are hatched for 15 days to 18 days, oyster shell and cement block substrata are placed into the ponds, and juvenile mollusk is obtained after the eyespot larvas adheres to the oyster shell and cement block substrata for four days to six days; finally, the juvenile mollusk adhering to the oyster shell and cement block substrata is moved into outdoor ponds to be bred, the juvenile mollusk is transported to a sea area in the south to be bred in an overland mode at lower temperature after the juvenile mollusk is 5 mm-10 mm long. By the adoption of the method suitable for artificial breeding of CrassOstrea hongkongensis in the sea area in the north, the problems that the rate of fertilization for artificial breeding of the CrassOstrea hongkongensis in the south is low and the survival rate of the larvas is low are solved.

The invention discloses crassostrea hongkongensis antimicrobial peptide sa-lectin as well as a recombinant expression method and application thereof. The gene coding sequence of antimicrobial peptide sa-lectin is shown in SEQ ID NO: 1, in which 156 amino acids are coded, and is shown in SEQ ID NO: 2, in which 16 amino acids at the end N are signal peptide sequences. According to the crassostrea hongkongensis antimicrobial peptide sa-lectin, a prokaryotic expression vector for removing the signal peptide sequence is constructed, so that sa-lectin recombined proteins are expressed and purified and can suppress growth of various types of gram-positive bacteria and gram-negative bacteria; furthermore, by the combination of sialic acid, an important effect is achieved in immune defense of shellfish.

The invention discloses microsatellite primers for identifying crassostrea hongkongensis, crassostrea ariakensis and crassostrea gigas and a hybrid thereof and an identification method.The microsatellite primers are F:5'-CGACTGGTGGGAGTTTCTGAC-3' and R:5'-GCCGCTTCTATCTCCTTTGC-3'.Due to the fact that the external morphology of oysters changes greatly along with different living environments, identification is usually difficult only according to morphological characteristics; the oysters in the larval stage are more difficult to distinguish according to the morphological characteristics.By means of the microsatellite primers and the identification method, crassostrea hongkongensis, crassostrea ariakensis and crassostrea gigas individuals can be identified rapidly and accurately, and identification of the hybrid individual of the crassostrea hongkongensis, crassostrea ariakensis and crassostrea gigas individuals has the potential application value.Meanwhile, the microsatellite primers for identifying the crassostrea hongkongensis, the crassostrea ariakensis and the crassostrea gigas and the hybrid thereof and the identification method have the advantages of being simple in method, visual, accurate and effective in result, free of influences of environment and growth stage, low in cost and the like.

The invention discloses a seedling clamping type crassostrea hongkongensis middle incubation method. The method comprises the following steps that (1) seedling clamping ropes are manufactured, wherein plastic ropes are cut, the length of the plastic ropes is 1.8-3 m, the diameter of the plastic ropes is 1.5-2.4 cm, each plastic rope is formed by two small ropes in a twisting mode, twist knots are arranged on the plastic ropes every 1.0-1.5 cm, the two ends of the plastic ropes are knotted, a 0.2-0.5 kg cement block is fixed at the tail end of each plastic rope to be used as a counterweight block, and manufacturing of the seedling clamping ropes is completed; (2) seedlings are clamped, wherein when young shells on a sheet-shaped plastic attaching piece are 2-10 mm in height, the twist knots in the step (1) are twisted off, fixing bases are clamped in the twist knots and fixed, and each seedling clamping rope is fixedly provided with 10-15 fixing bases; (3) middle incubation is conducted, wherein each seedling clamping rope with seedlings well clamped in the step (2) is placed in an ecological soil pool or a natural sea area, and the seedling clamping ropes are hung on a floating object for conducting middle incubation. The seedling clamping type crassostrea hongkongensis middle incubation method has the advantages that operation is convenient, time and labor are saved, the corrosion resistance capacity is high, the yield is large, and the benefits are high.

The invention discloses a time point quantification treatment method for producing a Crassostrea hongkongensis all-triploid. The time point quantification treatment method in which a fertilized egg development stage is taken as a biological indicator is created through the technical links of collection of single fertilized eggs, timely treatment of the fertilized eggs, detection of larva ploidy and the like, so that a 100-percent Crassostrea hongkongensis all-triploid can be obtained. Through optimization and pairing of the single fertilized eggs, the defect of poor mixed fertilized egg synchronicity in a conventional Crassostrea hongkongensis triploid induction process is overcome effectively, and zygotes with high synchronicity are obtained. A conventional fixed thinking mode in which a time point when 30-50 percent of a first polar body occurs is taken as a treatment time point and the optimal time interval is 20 minutes is overturned. Instead, the fertilized egg development stage is taken as the biological indicator, and A+1 / 3B and B+C are taken as a treatment time point and a treatment time interval respectively, so that the influences of external environmental conditions such as temperature on treatment are avoided, and the Crassostrea hongkongensis all-triploid can be induced successfully.

The invention discloses a non-damage preparation method for single crassostrea hongkongensis. The preparation method comprises the following steps: carrying out seedling collection by adopting a crassostrea angulata left shell as an adhering substrate, enabling seedlings of the crassostrea hongkongensis to be adhered onto the adhering substrate, and then carrying out seedling cultivation till the seedlings grow to be 30-60mm, so as to obtain adhering substrate combined clustered seedlings; placing the adhering substrate combined clustered seedlings into a culture cage, transferring the culture cage with the seedlings into a water body substrate-removing environment of which the salinity is 3-9 ppt, and the pH value is 6.00-6.75, and obtaining the single crassostrea hongkongensis after the crassostrea angulata left shell is molten. According to the invention, as the crassostrea hongkongensis has the characteristic of being capable of adapting to extreme environments with low salinity, low pH value and the like, the crassostrea angulata left shell is adopted as the adhering substrate, when the seedlings of the crassostrea hongkongensis grow to be 30-60 mm, the seedlings and the adhering substrate are placed in the culture cage, the culture cage is transferred into the environment with low salinity and low pH value, the cultivation is carried out for 15-30d, the adhering substrate is spontaneously and gradually molten through the ocean acidification reaction between slight acid brackish water and the crassostrea angulata left shell, the single crassostrea hongkongensis is prepared and scattered in the culture cage, and then, the single crassostrea hongkongensis is transferred to a proper culture area for continuing the cultivation.

The invention dislcoses a method for long-term storage of crassostrea hongkongensis eyespot larvas by critical low temperatures before the freezing point. The method comprises the steps that a, the larvas are stored in a refrigeration mode, the concentrated crassostrea hongkongensis eyespot larvas are collected and are packaged and bundled into packages by bolting-silk meshes which is wetted by seawater, and then the packages are wrapped with sponges wetted by seawater and are placed at the temperature of 0-1 DEG C for refrigeration; b, according to the refrigerated eyespot larvas in the step a, seawater is sprayed on the sponges, and the eyespot larvas are made to keep wet; c, the larvas are unfrozen, the refrigerated eyespot larvas in the step b are taken out from the low temperature environments and are wetted by normal-temperature seawater, then the larvas are put in a container and evenly stirred, and the eyespot larvas can be obtained after the larvas are completely restored to be at the normal temperature. The eyespot larvas stored in the method almost do not suffer from frostbite, the seedling collecting efficiency can be effectively improved, and 90% and more of the eyespot larvas can achieve metamorphosis. The method provides a theory and practice foundation for studying cryobiology of the crassostrea hongkongensis larvas, dry larvas transportation, large-scale seedling synchronous production and the like. The method for long-term storage of the crassostrea hongkongensis eyespot larvas by the critical low temperatures before the freezing point has the advantages that operation is simple and convenient, and the practicability is high.

The invention discloses a breeding method of a new thin-shell crassostrea hongkongensis variety. Crassostrea hongkongensis are different in shell thickness due to different breeding regions and are even of polymorphic biological characteristics of shell thickness in the same breeding regions, and according to the principle of negative correlation of the shell thickness and the dressing percentage, quantitative and economic synergic selection is realized by technical means of primary selection and secondary selection of shell thickness, self-breeding of population, oriental pure-breeding and the like in different breeding regions, and the new thin-shell crassostrea hongkongensis variety which is high in production and quality and stably genetic is obtained. Dressing percentage of 13-17% in conventional bred oysters is improved to 22-25%, and is improved by 30-90%, and firm practice foundation is created for improving yield of the breeding industry of the crassostrea hongkongensis. The breeding method has the advantages of simplicity and convenience in operation, high practicality, proneness to popularization and the like.

The invention discloses a cement knot type Crassostrea hongkongensis fixation base manufacture and application method. The invention develops a light corrosion-resistant cement knot type Crassostrea hongkongensis fixation base capable of saving labor force resource by the technical means, such as knot tying, cement rolling, fixation base manufacture, fixation base processing and fixation base application. According to the invention, the metamorphosis rate of the young Crassostrea hongkongensis is improved, the juvenile mollusk growth speed is quickened, the culture period is shortened, and the labor force cost is greatly reduced. The cement knot type Crassostrea hongkongensis fixation base is different from the traditional Crassostrea hongkongensis fixation base, such as heavy cement pats (blocks and bars), and shells which are easy to be subjected to acid corrosion by saline water and fresh water, and the cement knot type Crassostrea hongkongensis fixation base has the advantages of low cost, convenience in operation, time and labor saving, easiness in popularizing and the like..

The invention discloses a crassostrea hongkongensis LysM protein having a bacteriostatic activity, and an encoding gene and an application of the crassostrea hongkongensis LysM protein. The LysM protein is separated from crassostrea hongkongensis for the first time, PGN is effectively recognized, and the LysM protein has an obvious bacteriostatic effect on the growth of alginolyticus, staphylococcus haemolyticus, staphylococcus aureus and escherichia coli. The LysM protein plays important roles in immunological recognition of the crassostrea hongkongensis and the antibacterial activity, provides evidences for developing broad-spectrum antibiotics and screening an immunopotentiator, and also provides an application guide for the disease resistance and breeding of the shellfish.

The invention provides a sectional type efficient breeding method for crassostrea hongkongensis. The sectional type efficient breeding method for crassostrea hongkongensis comprises processes of juvenile collecting, medium juvenile culture, juvenile protection and fattening. A breeding mode of collecting juveniles by mattress sinking in a low-salt sea area in summers, carrying out rapid medium juvenile culture in a high-salt sea area in autumns and winters, protecting juveniles in the low-salt sea area in springs and rapidly fattening in high-salt sea area in autumns and winters is adopted, and fat crassostrea hongkongensis which reaches marketing specifications is obtained rapidly. Risks of death in springs in an area of collecting juveniles by mattress sinking are avoided, and the rate of survival of juvenile protection is increased. After a rapid juvenile culture stage in the high-salt sea area, juveniles at the age of 8 months can be the same as the original juveniles at the age of2 years, and the culture time of medium juvenile crassostrea hongkongensis is shortened by 16 months. The breeding period is shortened, the rate of survival of the juveniles is increased, the growthspecification of the crassostrea hongkongensis is large, and the economic benefit is good.

The invention discloses a method for cultivating a new crassostrea hongkongensis triploid rapid growth strain by using a backcross breeding technology. The method comprises the technical links of Crassostrea hongkongensis and Crassostrea rivularis diploid rapid growth line breeding, Crassostrea rivularis triploid induction line construction, Crassostrea rivularis tetraploid rapid growth line construction, hybrid Crassostrea rivularis diploid rapid growth line construction, backcross triploid rapid growth line cultivation and the like. The Crassostrea hongkongensis backcross triploid rapid growth new strain with faster growth and significantly improved survival rate in a high-salt region is obtained by backcrossing the Crassostrea hongkongensis tetraploid rapid growth line and the Crassostrea hongkongensis hybrid diploid rapid growth line and hybridizing different ploidy. Moreover, the new strain has the triploid advantages of extremely poor fertility, good taste and quality, capability of being marketed all year round, stable 100% triploid rate and the like, also has the advantages of pure crassostrea hongkongensis of white meat, tough meat, thick shell and the like, and is an excellent new strain suitable for commercial breeding in South China.

The invention discloses primers and a screening method of crassostrea hongkongensis triplet-base repetition microsatellite markers. By means of the screening method, a large quantity of crassostrea hongkongensis triplet-base repetition microsatellite markers can be obtained in a short time, and then an effective tool is provided for population genetic structure analyzing and molecular mark assisted breeding of the markers. According to the primers, amplification products of the primers have the good polymorphism, and the PCR amplification products are stable and can be applied to analysis on crassostrea hongkongensis diversity and a population genetic structure.

The invention discloses a method for producing allotetraploid by hybridizing crassostrea angulata diploid and crassostrea hongkongensis triploid. According to the method, the crassostrea hongkongensistriploid is used as a female parent, the full weight is used as a target, the crassostrea angulata diploid is preferably selected as a male parent, and the oyster allotetraploid is produced through drug-induced hybridization. The oyster allotetraploid obtained by the method has the characteristics of euryhalinity and high temperature resistance, is suitable for breeding in low-salt areas such asestuary, and the oyster triploid produced by hybridizing the oyster allotetraploid with the common crassostrea angulata or crassostrea hongkongensis diploid is suitable for aquaculture in sea areas with different salt contents in China , and has wide popularization and application prospects.

The invention discloses a step-by-step fresh water soaking method for preventing crassostrea hongkongensis from mass mortality. The method includes the steps: a, transferring to a low-salt environment, regularly checking growth and development situations of crassostrea hongkongensis, pulling a floating mat of the crassostrea hongkongensis from a sea area into a low-salinity area with transfer inter-area salinity gradient difference more than or equal to 8ppt once more than 8% of the crassostrea hongkongensis dies, continuing cultivation and observing whether mortality of the crassostrea hongkongensis is inhibited or not; b, performing ultralow-salinity treatment, when the crassostrea hongkongensis after being treated at the step a still continues to die, transferring the crassostrea hongkongensis treated at the step a to an ultralow-salinity environment area with salinity less than or equal to 5ppt or even a pure fresh water area to be cultivated. Survival rate of the crassostrea hongkongensis treated by the method is above 90% generally, and is increased by 30-90% as compared with that of a traditional cultivation technique; yield can be increased by 150,000-300,000 tons, output value is increased by 1 billion to 2 billion RMB, and a solid practice foundation is laid for sustainable and healthy development of the crassostrea hongkongensis cultivation industry. The method has the advantages of high operability, significant effect, easiness in popularization and the like.

The invention discloses a crassostrea hongkongensis BPI gene, an encoded protein and a cloning method therefor, and a construction method for a crassostrea hongkongensis BPI genetic engineering strainand application of a recombinant protein thereof. The sequence of the crassostrea hongkongensis BPI gene is shown by ESQ ID NO:1, and the cloning method comprises designing a specific primer amplified gen according to a conserved sequence isogenous to the BPI gene; a sequence of the crassostrea hongkongensis BPI protein is shown by SEQIDNO.2; primers containing an Ndel locus and an Xbal locus areconfigured to amplify a crassostrea hongkongensis BPI mature protein; and a target gene obtained by cloning is inserted into a carrier to obtain a recombinant plasmid, inducible expression is conducted on the recombinant plasmid, and the genetic engineering strain is obtained through purification and renaturation. The method has the advantage of having an obvious bactericidal effect on vibrio alginolyticus and vibrio parahaemolyticus.

The invention discloses a method for obtaining crassostrea hongkongensis monomers through medicine induction. The method comprises steps of cultivating larvae, screening larvae at foot and face growing stage, performing medicine induction, performing cultivation with descending flows, performing juvenile mollusk management and the like. Cultivation is performed according to the crassostrea hongkongensis fry breeding technical regulations, larvae are screened to obtain larvae at foot and face growing stage of which the specifications are 300-350[mu]m, then the larvae at the foot and face growing stage are induced with inducing liquid, then cultivation is performed with the descending flows, and after 8-10 days, crassostrea hongkongensis monomer juvenile mollusks of which the shell height is1-2mm can be obtained. According to the method disclosed by the invention, through medicine stimulation, the larva at the foot and face growing stage can be smoothly induced, low distortion ratio andsecondary hurt during base removal are avoided, the non-adherence distortion ratio of the larvae is greatly increased, the distortion ratio of the larvae is 60% or above, the larvae which are not successfully induced can be recovered for continuous cultivation, after feet appear, induction is performed anew, and the problems that crassostrea hongkongensis fries are easy to damage, the survival rate is low and the yield in unit water body is low in breeding process can be solved.

The invention discloses a method for inducing efficient attachment metamorphosis of Crassostrea hongkongensis. The method comprises the following steps: (1) after Crassostrea hongkongensis larvae arecultured to a certain specification, starting to decrease salinity of a culture water body in each day, and carrying out conventional feeding management; (2) moving a cleaned and disinfected attachment medium into an attachment metamorphosis pond, filling the pond with water for the first time, carrying out draining, filling the pond with micron-level filtered seawater, adding EDTA and air heads;(3) after detecting that the ratio of eyespot larvae is over a half by virtue of a microscope, pouring the larvae into the pond, carrying out sieving and screening, transferring big larvae into the attachment metamorphosis pond in the step (2), uniformly scattering ZnCl2 induction liquid into the pond, and carrying out air inflation and uniform mixing; and (4) after the attachment metamorphosis ofthe eyespot larvae is finished, draining the water, injecting the micron-level filtered seawater into the pond again, carrying out decrement feeding in the first day, carrying out increment feeding in the second day, and carrying out doubled feeding management after 72h. The method is simple, convenient and feasible, is suitable for large-scale production, application and popularization and has obvious promotion effects to other oysters.

The invention discloses a breeding method for new line of backcross Crassostrea hongkongensis and hybrid gigas. According to the technical links of sperm and ovum preparation, fertilization incubation, filial generation breeding and the like, the characteristic of hybrid F1 male sterile is overcome; a male individual which has the characteristics of a hybrid F1 and is completely fertile in a hybrid F2 is used as a sperm source and is fertilized with a Crassostrea hongkongensis ovum to successfully breed the new line of backcross Crassostrea hongkongensis and hybrid gigas. The breeding method disclosed by the invention is different from the characteristic in the traditional backcrossing that the hybrid F1 is used for backcrossing with a parent, and the hybrid which has the characteristics of a hybrid F1 and is completely fertile in a hybrid F2 is used for backcrossing with the Crassostrea hongkongensis to breed a new line. The new line obtained with the breeding method disclosed by the invention has the advantages of wide temperature applicable range, strong high salinity resistance, obvious growth and survival advantage, simpleness in operation, strong practicality and easiness in popularization, can be bred from north to south, the gigas breeding area is obviously enlarged, and the yield of the new line is improved by 40-150% if being compared with that of the Crassostrea hongkongensis.

The invention discloses a manufacture and application method for a cement shell composite type Crassostrea hongkongensis fixation base suitable for Crassostrea hongkongensis to cultivate from collecting fries. According to the manufacture and application method disclosed by the invention, one set of cement shell composite type Crassostrea hongkongensis fixation base with high fry collection efficiency and strong resistance to corrosion is developed by the technical means of material preparation, fixation base manufacture, fixation base processing, fixation base fry collection and the like. According to the manufacture and application method disclosed by the invention, the fry collection area of the fixation base is increased, the metamorphosis rate of the young Crassostrea hongkongensis is improved, the juvenile mollusk growth speed is quickened, and the corrosion function in the saline water and fresh water environment can be effectively resisted. The manufacture and application method is not only suitable for indoor artificial fry collection but also suitable for outdoor natural fry collection. The cement shell composite type Crassostrea hongkongensis fixation base disclosed by the invention is different from the traditional Crassostrea hongkongensis fixation base, such as heavy cement pats (blocks and bars), and shells which are easy to be subjected to acid corrosion by saline water and fresh water, and the cement shell composite type Crassostrea hongkongensis fixation base combines the advantages of the heavy cement pats and the shells. The manufacture and application method has the advantages of high colony-forming efficiency, low cost, easiness in operation, convenience in popularization and the like.

The invention discloses an autophagy protein beclin-1 as well as a coding gene and application thereof. The amino acid sequence of the autophagy protein beclin-1 is as shown in SEQ ID NO. 2. A Beclin-1 gene is obtained through identification from crassostrea hongkongensis for the first time, formation of autophagosome can be inhibited by interfering expression of Beclin-1, an NF-kappaB signal channel is inhibited, and infection of vibrio alginolyticus and staphylococcus haemolyticus can be quickly responded. The Beclin-1 plays an important role in immune defense of the crassostrea hongkongensis, and provides application guidance for disease control and disease-resistant breeding of the crassostrea hongkongensis and other mariculture shellfishes.