CN116548386A - Efficient preparation and detection method and application of super-male grass carp - Google Patents
Efficient preparation and detection method and application of super-male grass carp Download PDFInfo
- Publication number
- CN116548386A CN116548386A CN202310452423.6A CN202310452423A CN116548386A CN 116548386 A CN116548386 A CN 116548386A CN 202310452423 A CN202310452423 A CN 202310452423A CN 116548386 A CN116548386 A CN 116548386A
- Authority
- CN
- China
- Prior art keywords
- grass carp
- male
- super
- development
- eggs
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 241000252230 Ctenopharyngodon idella Species 0.000 title claims abstract description 61
- 238000001514 detection method Methods 0.000 title claims abstract description 16
- 238000002360 preparation method Methods 0.000 title claims abstract description 7
- 238000000034 method Methods 0.000 claims abstract description 25
- 238000011161 development Methods 0.000 claims abstract description 24
- 230000018109 developmental process Effects 0.000 claims abstract description 24
- 238000009395 breeding Methods 0.000 claims abstract description 4
- 230000001488 breeding effect Effects 0.000 claims abstract description 4
- 235000013601 eggs Nutrition 0.000 claims description 17
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 15
- 241000251468 Actinopterygii Species 0.000 claims description 12
- 230000012447 hatching Effects 0.000 claims description 7
- 230000035939 shock Effects 0.000 claims description 6
- 230000004720 fertilization Effects 0.000 claims description 5
- 238000011534 incubation Methods 0.000 claims description 5
- 238000002635 electroconvulsive therapy Methods 0.000 claims description 4
- 210000000349 chromosome Anatomy 0.000 abstract description 6
- 238000011160 research Methods 0.000 abstract description 4
- 230000020509 sex determination Effects 0.000 abstract description 4
- 241001465754 Metazoa Species 0.000 abstract description 3
- 230000004069 differentiation Effects 0.000 abstract description 2
- 238000012214 genetic breeding Methods 0.000 abstract description 2
- 230000008569 process Effects 0.000 abstract description 2
- 239000003147 molecular marker Substances 0.000 abstract 1
- 238000000746 purification Methods 0.000 abstract 1
- 239000003550 marker Substances 0.000 description 8
- 108020004414 DNA Proteins 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 210000004940 nucleus Anatomy 0.000 description 6
- 210000004681 ovum Anatomy 0.000 description 6
- 238000011282 treatment Methods 0.000 description 6
- 238000006243 chemical reaction Methods 0.000 description 5
- 210000001766 X chromosome Anatomy 0.000 description 4
- 210000002593 Y chromosome Anatomy 0.000 description 4
- 230000003321 amplification Effects 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 230000006698 induction Effects 0.000 description 4
- 238000003199 nucleic acid amplification method Methods 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 102000002322 Egg Proteins Human genes 0.000 description 3
- 108010000912 Egg Proteins Proteins 0.000 description 3
- 108091028043 Nucleic acid sequence Proteins 0.000 description 3
- 238000004090 dissolution Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 238000012163 sequencing technique Methods 0.000 description 3
- 238000012408 PCR amplification Methods 0.000 description 2
- 230000013020 embryo development Effects 0.000 description 2
- 230000004907 flux Effects 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 238000002844 melting Methods 0.000 description 2
- 230000008018 melting Effects 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- QLGKMLRGKPKGKI-QWQCQAQZSA-N (2s)-n-[(2s)-1-[[(2s)-1-[[(2s)-1-[[(2s)-1-[[(2r)-1-[[(2s)-1-[[(2s)-5-(diaminomethylideneamino)-1-[(2s)-2-(ethylcarbamoyl)pyrrolidin-1-yl]-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]amino]-1-oxopropan-2-yl]amino]-3-(4-hydroxyphenyl)-1-oxopropan-2-yl Chemical compound CCNC(=O)[C@@H]1CCCN1C(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](C)NC(=O)[C@@H](NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H]1NC(=O)CC1)CC1=CC=C(O)C=C1 QLGKMLRGKPKGKI-QWQCQAQZSA-N 0.000 description 1
- 241000252185 Cobitidae Species 0.000 description 1
- 241000252233 Cyprinus carpio Species 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 108010044467 Isoenzymes Proteins 0.000 description 1
- 241001327682 Oncorhynchus mykiss irideus Species 0.000 description 1
- 241000276703 Oreochromis niloticus Species 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 210000002257 embryonic structure Anatomy 0.000 description 1
- 238000012252 genetic analysis Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 238000003205 genotyping method Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 230000009027 insemination Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000001161 mammalian embryo Anatomy 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000001823 molecular biology technique Methods 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 210000000006 pectoral fin Anatomy 0.000 description 1
- 238000000053 physical method Methods 0.000 description 1
- 238000003976 plant breeding Methods 0.000 description 1
- 238000012257 pre-denaturation Methods 0.000 description 1
- 239000003761 preservation solution Substances 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention belongs to the technical field of aquatic animal genetic breeding, and discloses a high-efficiency preparation and detection method and application of a super-male grass carp, wherein the super-male grass carp of YY-type chromosome can be cultivated by the method, and meanwhile, quick HRM typing is performed by utilizing the development molecular marker of the invention. The YY super-male grass carp cultivated by the method provided by the invention can be applied to rapid purification of germplasm in the breeding process of the grass carp, and can also be applied to basic researches such as sex determination and differentiation of the grass carp.
Description
Technical Field
The invention relates to the technical field of aquatic animal genetic breeding, in particular to a high-efficiency preparation and detection method and application of a super-male grass carp, and the super-male grass carp with YY-type chromosomes can be cultivated and rapidly screened by adopting the method.
Background
Male nuclear development refers to the way in which genetically inactivated ova fertilize with sperm, producing offspring containing only male genetic material. At present, no report on natural male nucleus development of fishes exists, but the natural male nucleus development can be realized through artificial induction, and the common practice is to treat ova by physical methods such as ultraviolet rays, X rays, gamma rays and the like, so that the ova are inactivated and then participate in fertilization, and then the chromosomes of sperm nuclei are doubled by treatments such as cold, heat, high pressure and the like at proper time, so that the sperm nuclei are developed into diploid which is completely male parent characters. Because the male nuclear development individuals are highly homozygous, the male nuclear development individuals can not only generate super male and unisexual offspring, but also be used for quickly establishing pure lines, distinguishing sex determination mechanisms, protecting endangered species and the like, and thus, the male nuclear development individuals have very important significance for the research of the male nuclear development individuals.
However, compared with the artificial induction of gynogenesis, the survival rate of the individuals with the artificial induction of the gynogenesis is lower, the present androgenetic development is applied more in plant breeding, and the preparation of super-male fish usually needs cold, hot or high-pressure treatment in the embryo development period, but the starting time of the treatment is not definite, and an accurate starting time is difficult to find. There are few studies on fish among aquatic animals, and the fish subjected to the current male nuclear development study include rainbow trout, loach, carp, nile tilapia, and the like.
Since the handling of artificially inactivated egg chromosomes is not one hundred percent successful, there must be evidence that the egg checking embryo does not provide genetic material, which involves the problem of identifying a male-developing diploid with a diploid that would result from normal fertilization. Early identification methods include morphological genetic markers, chromosome typing, isozymes, etc., but are mostly complex and cannot be accurately identified. With the development of molecular biology, DNA fingerprint, RAPD marker and other technical means appear, so that the DNA markers of male and female parents and offspring of male nuclear development can be analyzed, and the source of offspring genetic material is ascertained, thus being a more accurate analysis method in male nuclear development. Some domestic and foreign researches verify the correctness of the androgenetic pathway through molecular biology techniques such as RAPD, and the corresponding method can be used for identifying the androgenetic individuals in turn, such as SNP, indel markers and the like which are popular at present. HRM (high resolution melting curve) analysis technology is a new genetic analysis method for mutation scanning and genotyping which has been raised in recent years. Based on the efficient and robust PCR technology, the HRM does not need a sequence specific probe, directly runs high-resolution melting after the PCR is finished, and has the advantages of high sensitivity, high flux, low cost and the like. However, the HRM also has high requirements on the length of the PCR product (less than 100 bp), and the number of the difference bases in 100bp is more than or equal to 8 in order to ensure good resolution. The molecular markers of grass carp gender which are developed at present only contain sporadic SNP markers, are far away and are not suitable for the HRM analysis technology.
In the invention, after ultraviolet irradiation inactivation is carried out on the grass carp ovum, the grass carp ovum is inseminated with the grass carp sperm, and in any time point of 6-36min of development, the fertilized ovum can be taken out for cold shock treatment and then transferred into a hatching barrel for hatching, thus obtaining the super-male grass carp. Meanwhile, the applicant also carries out fine comparison on female and male genomes of the grass carp, discovers a grass carp X/Y chromosome specific marker suitable for the HRM analysis technology, and can successfully distinguish XX, XY and YY super-male grass carp by using the marker to identify the grass carp with male nucleus development.
The invention not only can generate super male grass carp and unisexual offspring to quickly establish the grass carp pure line, but also can judge the sex determination mechanism of the grass carp, and is used for basic researches such as sex determination and differentiation of the grass carp.
Disclosure of Invention
The invention aims to provide a method for efficiently preparing and detecting supermale grass carp, which is simple, convenient and quick.
The invention further aims at providing an application of the method for efficiently preparing and detecting the supermale grass carp.
In order to achieve the above object, the present invention adopts the following technical measures:
the efficient preparation and detection method of the super-male grass carp comprises the following steps:
and (3) after the grass carp eggs are inactivated, the grass carp eggs are incubated with grass carp sperms for fertilization, the grass carp eggs are transferred into a water body at 25 ℃ for development, the fertilized eggs are subjected to cold shock treatment at any time point in the development period, then are hatched, and after the hatching, the fish fries are subjected to HRM typing detection by adopting primers CIYSPE-F GGAGAGACTGACAGGTGTGTGAC and CIYSPE-R CCCCCAACGCTCGGAGGT.
In the above method, preferably, the period of development is in the range of 6-26min;
in the above method, preferably, the cold shock is performed in a water bath at 4-6deg.C for 8-16min;
in the method, preferably, the method is characterized in that the cold shock is carried out in a water bath at the temperature of 4 ℃ for 12min;
in the above method, the incubation water temperature is preferably 22-28deg.C, and most preferably 25deg.C.
The protection content of the invention also comprises: the method is applied to the breeding of the super-male grass carp; the application of the super-male grass carp prepared by the method in the establishment of grass carp pure line.
Compared with the prior art, the invention has the following advantages and effects:
there is no successful technique for preparing and identifying YY supermale grass carp, and cold, hot or high pressure treatment is usually required during embryo development, but the starting time of treatment is not definite, and it is difficult to find out an accurate starting time. The invention does not need to determine the time of the cold shock initiation of fertilized eggs of grass carp, and can induce the super-male grass carp to be obtained by carrying out water bath cold shock treatment on embryos at any time point within the time of 6-36min after fertilization.
In addition, in order to rapidly identify the sex of the grass carp by using the HRM technology, the invention obtains the male nucleus development YY grass carp and simultaneously screens and obtains a grass carp X/Y chromosome specific marker, wherein the marker comprises 3 SNP loci and 1 Indel locus, is suitable for the detection of the HRM technology, can rapidly identify the sex of the obtained grass carp, and accurately distinguishes three chromosome types of XX, XY and YY. The detection can be completed in 96-well plates, and 10 96-well plates, namely 960 samples can be completed in one day. Therefore, the YY super-male individual detection efficiency is high, the flux is large, the detection period is greatly shortened, and the cost caused by subsequent test cross verification is reduced.
Description of the drawings:
FIG. 1 shows the detection results of XX, XY and YY grass carp HRM using the primer pair provided by the invention;
the upper graph is a dissolution curve showing that the fluorescence intensity gradually decreases (Y axis) with increasing temperature (X axis), reflecting the effect of the temperature increase on the dissociation degree of the DNA double helix structure;
the lower graph shows the dissolution peak curve, showing the change in the derivative of fluorescence intensity value with temperature (Y axis) with increasing temperature (X axis), reflecting the inconsistency in the dissolution temperature (peak temperature) when DNA duplex of the amplification products of three XX, XY and YY dissociates in half.
Detailed Description
The technical scheme of the invention is a conventional scheme in the field unless specifically stated; the reagents or materials, unless otherwise specified, are commercially available.
Example 1:
induction parameters of grass carp YY supermale individuals were explored:
1) 1 day before artificial breeding of grass carp, female fish is injected with DOM (dosage/weight of 1 mg/Kg), LHRH-A2 (dosage/weight of 5 μg/Kg) and HCG (dosage/weight of 1000 IU/Kg) under pectoral fin, and male fish dosage is halved. The effect time is about 21h under the temperature control condition of 26 ℃.
2) About 500 grass carp eggs and 100mL hank's liquid are added into each glass dish with the diameter of 25cm, the glass dishes are placed on a shaking table (80 r/min), and two ultraviolet lamp tubes with the width of 15W (the vertical distance between the ultraviolet lamp and the culture dish is 15 cm) are irradiated for 30s-45s, so that female procaryotes are inactivated.
3) Grass carp sperm were collected at a ratio of 1: adding D-15 preservation solution according to the proportion of 10, mixing uniformly, carrying out wet insemination on the mixture with the eggs inactivated by ultraviolet irradiation in water with the temperature of 25 ℃ for 2min, and transferring the mixture to a water body with the temperature of 25 ℃ for development.
4) And respectively developing for 6min, 8min, 10min, 12min, 14min, 16min, 18min, 20min, 22 min, 24min, 26min and 36min, collecting part of fertilized eggs, treating in water at 4deg.C for 12min, and transferring to a 25 deg.C incubation barrel for incubation. The hatching rate of the cold shock at the time point is respectively 0.32 per mill, 0.30 per mill, 0.41 per mill, 0.38 per mill, 0.66 per mill, 0.87 per mill, 0.72 per mill, 1.29 per mill, 2.46 per mill, 2.64 per mill and 1.57 per mill. 0.29 per mill.
The formula for calculating the hatching rate is as follows: hatching tail seedlings/chilled fertilized eggs 1000%
5) And finally, from 200,000 induced fertilized eggs, about 200 seedlings are obtained after membrane emergence, all male nuclear development samples are collected when the fertilized eggs are cultured to 1 month of age, and the tail fin tissues are taken to extract genome DNA.
Grass carp X/Y chromosome specific marker screening:
1) 8 individuals of female and male grass carp in different water areas with determined sexes are selected for resequencing (PE 150), and the data amount is about 30G/tail. The female and male sequencing data were mixed into female and male mixing pools, respectively, and subjected to electronic cleavage treatment with a Kmer of 51bp starting from the six bases AC, AG, AT, GA, GC and GT. After redundancy removal, the frequency of each Kmer was obtained, kmers with frequency of 1 were removed, and the two were compared to obtain a male-specific Kmer.
2) Male specific Kmers were assembled using Velvet software to obtain Y-contig. Using Y-contig as reference, kmer generated by the female and male mix pools was used as query, and default alignment was performed using Boutai2 software. Sites with a difference between SNP and Indel of greater than or equal to 10bp in the female and male pool Kmers were screened in the 100bp region.
3) Further, in sequencing reads of 16 tail females and 16 tail males in different water areas, the regions are completely aligned (-all), and conserved differential SNP and Indel sites are found, so that sex distinction can be realized.
4) PCR detection primers were designed on both sides of SNP and Indel sites with large sequence differences using IGV software visualization Manual check. Finally, CIYSPE sequences containing SNP and Indel marker sequences in male Y chromosomes are obtained, the nucleotide sequences are shown as SEQ ID NO.1, the corresponding sequences in X chromosomes in females are shown as SEQ ID NO.2, and compared with the marker sequences in males, the CIYSPE sequences have 3 SNP differences (G- & gt T, C- & gt A and A- & gt G) and are inserted with TGTGTGTGTGA sequences, so that the super-male grass carp identification can be carried out by designing primers aiming at the nucleotide sequences which are different and contain the sequences of SEQ ID NO.1 and SEQ ID NO.2 in grass carp.
HRM detection of male-nuclear developed fries:
after amplification using primers (CIYSPE-F: GGAGAGACTGACAGGTGTGTGAC; CIYSPE-R: CCCCCAAC GCTCGGAGGT), typing was performed on an HRM-apparatus, and the typing results are shown in FIG. 1. And 81 fish fries in the 200 fish fries are super-male grass carp, 10 fish fries are randomly selected from the 81 super-male grass carp to carry out genome re-sequencing, and the two fish fries are compared with grass carp reference sequences to form types, namely YY-type grass carp, and no false positive is generated.
Example 2:
the application process of the sequences shown in SEQ ID NO.1 and SEQ ID NO.2 in the identification of the supermale of grass carp comprises the following steps:
1) Primers were designed for the difference nucleotide sequences between SEQ ID NO.1 and SEQ ID NO. 2:
CIYSPE-F:GGAGAGACTGACAGGTGTGTGAC
CIYSPE-R:CCCCCAACGCTCGGAGGT
2) Genomic DNA extraction:
mu.l of 5% strength chelex100 was added to a 96-well plate equipped with tail fins, digested at 58℃for 1 hour, boiled at 100℃for 8 minutes, centrifuged at 4000 rpm for 5 minutes, and 1. Mu.l of the supernatant was used as a template for PCR reaction.
3) And (3) PCR amplification:
a reaction system was prepared in a 96-well plate, and the reaction system was 10 XBuffer (Mg2+plus) 1. Mu.l; LC Green 1 μl; dNTP (10 mM each) 0.2uL; CIYSPE-F (10 mM) 0.2uL; CIYSPE-R (10 mM) 0.2uL; taq (5U/uL) 0.1uL; 1 μl of template DNA was finally supplemented with ddH2O to 10 μl. The PCR reaction conditions are 94 ℃ for pre-denaturation for 4min; denaturation at 94℃for 30s, annealing at 55℃for 30s, elongation at 72℃for 25s,40 cycles; final extension at 72℃for 5min; preserving at 4 ℃.
The amplified specific sequences in the supermale are:
GGAGAGACTGACAGGTGTGTGACACGTGATCTACAGGACAGTGAGGCCACAATGAGATGAGATCGTGTGTGTACCTCCGAGCGTTGGGGG；
specific sequences amplified in female individuals are:
GGAGAGACTGACAGGTGTGTGACACGTGATCTACATGACAGTGAGGACACAATGAGGTGAGATCTGTGTGAGTGTGTGTACCTCCGAGCGTTGGGGG；
the specific sequences amplified in the male individuals are:
GGAGAGACTGACAGGTGTGTGACACGTGATCTACAGGACAGTGAGGCCACAATGAGATGAGATCGTGTGTGTACCTCCGAGCGTTGGGGG；
and GGAGAGACTGACAGGTGTGTGACACGTGATCTACATGACAGTGAGGACACAATGAGGTGAGATCTGTGTGAGTGTGTGTACCTCCGAGCGTTGGGGG.
For the PCR amplification products described above, detection of the amplification products can be performed using protocols conventional in the art.
In the invention, the amplification result is detected by using HRM, after the PCR reaction is finished, a 96-well plate is directly put into a LightScanner 96 instrument plate groove for HRM scanning, the initial scanning temperature is set at 68 ℃, the end scanning temperature is set at 94 ℃, and the Start Run is clicked to Start scanning; after the scanning is finished, a separation curve is obtained.
The curve can completely separate female individuals, male individuals and supermale individuals of grass carp.
Claims (8)
1. The efficient preparation and detection method of the super-male grass carp comprises the following steps:
and (3) after the grass carp eggs are inactivated, the grass carp eggs are incubated with grass carp sperms for fertilization, the grass carp eggs are transferred into a water body at 25 ℃ for development, the fertilized eggs are subjected to cold shock treatment at any time point in the development period, then are hatched, and after the hatching, the fish fries are subjected to HRM typing detection by adopting primers CIYSPE-F GGAGAGACTGACAGGTGTGTGAC and CIYSPE-R CCCCCAACGCTCGGAGGT.
2. The method according to claim 1, characterized in that: the time range of the development period is 6-26min.
3. The method according to claim 1, characterized in that: the cold shock is performed in water bath at 4-6deg.C for 8-16min.
4. A method according to claim 3, characterized in that: the cold shock is carried out by treating in water bath at 4 ℃ for 12min.
5. The method according to claim 1, characterized in that: the incubation water temperature is 22-28 ℃.
6. The method according to claim 5, wherein: the incubation water temperature was 25 ℃.
7. Use of the method of claim 1 in the breeding of hyperactive grass carp.
8. Use of the method of claim 1 for the establishment of grass carp pure lines.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310452423.6A CN116548386B (en) | 2023-04-25 | Efficient preparation and detection method and application of super-male grass carp |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310452423.6A CN116548386B (en) | 2023-04-25 | Efficient preparation and detection method and application of super-male grass carp |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN116548386A true CN116548386A (en) | 2023-08-08 |
| CN116548386B CN116548386B (en) | 2024-05-10 |
Family
ID=
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4697546A (en) * | 1986-05-30 | 1987-10-06 | Purdue Research Foundation | Method for production of tetraploid channel catfish |
| CN1340298A (en) * | 2000-08-25 | 2002-03-20 | 武汉康祥科技发展有限公司 | Technique for creating super-male fish reproduction system |
| CN107079844A (en) * | 2017-05-08 | 2017-08-22 | 中国科学院水生生物研究所 | A kind of preparation method of the female carp of supermale carp and Genetic male and physiology |
| CN109554486A (en) * | 2018-12-30 | 2019-04-02 | 中国水产科学研究院珠江水产研究所 | SNP marker relevant to grass carp character and its application |
| CN113930525A (en) * | 2021-11-19 | 2022-01-14 | 中国科学院水生生物研究所 | Specific sequence for snakehead sex identification and application |
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4697546A (en) * | 1986-05-30 | 1987-10-06 | Purdue Research Foundation | Method for production of tetraploid channel catfish |
| CN1340298A (en) * | 2000-08-25 | 2002-03-20 | 武汉康祥科技发展有限公司 | Technique for creating super-male fish reproduction system |
| CN107079844A (en) * | 2017-05-08 | 2017-08-22 | 中国科学院水生生物研究所 | A kind of preparation method of the female carp of supermale carp and Genetic male and physiology |
| CN109554486A (en) * | 2018-12-30 | 2019-04-02 | 中国水产科学研究院珠江水产研究所 | SNP marker relevant to grass carp character and its application |
| CN113930525A (en) * | 2021-11-19 | 2022-01-14 | 中国科学院水生生物研究所 | Specific sequence for snakehead sex identification and application |
Non-Patent Citations (1)
| Title |
|---|
| 赵振山, 吴清江, 高贵琴: "鱼类雄核发育的研究进展", 遗传, no. 02 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106381331B (en) | The relevant SNP marker of Growth of Grass Carps Ctenopharyngodon Idellus speed and its application | |
| CN106399293B (en) | High-efficiency induction and microsatellite molecular marker identification method for nile tilapia pseudofemale fish | |
| CN102134593A (en) | Gender-specific microsatellite marker for Cynoglossus semilaevis and application of same in identification of superfemale Cynoglossus semilaevis | |
| CN113881785B (en) | SNP locus primer combination for multi-character breeding of litopenaeus vannamei and application | |
| CN106591429B (en) | Method for screening sex reversal genetic control sites of cynoglossus semilaevis, kit and application | |
| CN109554486A (en) | SNP marker relevant to grass carp character and its application | |
| CN103798169A (en) | Method for quickly establishing improved breeding system of YY super-male pelteobagrus fulvidraco | |
| CN114657264B (en) | Clarias fuscus sex-specific molecular marker primer and application thereof | |
| CN111394445B (en) | Indel marker for sex identification of channa maculata and application thereof | |
| CN111378731B (en) | Sex-specific molecular marker primer for Trachinotus ovatus and application of sex-specific molecular marker primer | |
| CN101921849B (en) | Method for assisting to authenticate milk cows with different milk producing characteristics and special primer pairs used by same | |
| CN111560401A (en) | Molecular breeding method for thickening interpuscular spurs of erythroculter ilishaeformis and megalobrama amblycephala | |
| CN105441536B (en) | SNP marker for discriminating sex in paralichthys olivaceus | |
| CN116548386B (en) | Efficient preparation and detection method and application of super-male grass carp | |
| CN103409516A (en) | Identification method of channel catfish species | |
| CN113249492B (en) | SNP marker for evaluating pig eye muscle area and application method thereof | |
| CN116548386A (en) | Efficient preparation and detection method and application of super-male grass carp | |
| CN104988240A (en) | Method for identifying swarm royal jelly high yield character with SNP mark rs16287910 | |
| CN115044682A (en) | Molecular marker related to growth traits of Hu sheep, detection method and application thereof | |
| Chakraborty et al. | Applied molecular cloning: present and future for aquaculture | |
| CN114574594A (en) | California perch sex specific SNP molecular marker primer and application thereof | |
| CN113684282A (en) | Method for detecting pig hairless character based on EDAR gene expression level and application thereof | |
| KR102000878B1 (en) | Marker for male and female of fish identification and HRM-PCR analysis method | |
| CN108841971B (en) | Method for detecting cattle SH3PXD2B gene insertion/deletion marker | |
| CN101633936B (en) | Fish transgenosis breeding method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant |