CN105002267A - Penaeus japonicus molecule marking method and application - Google Patents
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Abstract
The invention relates to a penaeus japonicus molecule marking method and application. The penaeus japonicus molecule marking method comprises the steps of extracting a DNA genome, performing PCR amplification and product identification and is applied to diversity analysis of screened penaeus japonicus germplasm resources, genetic diversity analysis, parentage assignment, genetic research of molecular population, genetic map establishment, important economic character positioning, functional gene research and assisting of penaeus japonicus molecule genetic breeding or cultivation. By means of the penaeus japonicus molecule marking method, a genetic variation polymorphic map of a genetic marker gene locus of the penaeus japonicus can be rapidly obtained, the method is simple, convenient and quick, and a result can be visually observed. The penaeus japonicus molecule marking method is mainly applied to genetic marking, genealogical identification and genetic map establishment of a penaeus japonicus colony and is rapid in detection, low in cost and wider in application range.
Description
Technical field
The invention belongs to molecular biology DNA molecular Genetic Markers, be specifically related to marine economic animal japonicus molecule marking method and application.
Background technology
Japonicus has another name called colored shrimp, ring shrimp, flower tail shrimp, spot joint shrimp, car shrimp, and be extensively distributed in the Indian Ocean, the Pacific region, coastal on the south Jiangsu Province, China also have distribution.Japonicus have growth rapidly, disease resistance is strong, dried up long-time dead, being applicable to long-distance dry fortune active pin, is important fishery shrimps of earning foreign exchange.Japonicus is nutritious, containing protein be fish, egg, several times to tens times of milk; Also containing abundant mineral substance and the composition such as vitamin A, aminophylline such as iodine, potassium, magnesium, phosphorus, and its meat is the same with fish soft, easy to digest, the nutrient excellent product that the elderly that can yet be regarded as uses, and has benifit to healthy pole; To in poor health and need the people taken good care of also to be fabulous food after being ill.Therefore Fed Penaeus japonicus is for raiser being a fabulous income.
But the fast development of cultivation and the disease that produces makes japonicus resource decay rapidly, especially since the prawn disease outburst nineties, japonicus aquaculture is dropped sharply, and the popular disease-resistant strain breeding of prawn that makes of prawn disease becomes the problem necessarily solved.At present, domestic fishery genetic improvement is mainly based on unexpected mass incident, but the offspring that unexpected mass incident obtains, between its individuality, sibship is indefinite, inevitably causes the loss of inbreeding and genetic diversity in Breeding Process.Therefore, strain improvement needs to carry out with more accurate selection.These methods are badly in need of finding a kind of mark that effectively can identify different family, thus know the sibship of breeding population, understand pedigree information, effectively reduce inbreeding, and rationally utilize breeding population efficiently.
Chinese patent notification number CN 100532573C, August 26 2009 day for announcing, name is called the screening method of pacific oyster EST micro-satellite mark, this application case discloses a kind of screening method of pacific oyster EST micro-satellite mark, comprise the sequence of the ESTs of the Pacific oyster utilizing GeneBank to announce, micro-satellite retrieval software SSRhunter is utilized to carry out searching of microsatellite locus, micro-satellite segment 2-6 base repeating unit multiplicity being greater than to 5 carries out screening and separating, thus obtains the ESTs sequence containing micro-satellite repetition; Then the flanking sequence design primer at micro-satellite repetitive sequence two ends, and inspection optimization primer becomes microsatellite marker further; Finally comprehensive evaluation is carried out to the repeatability of microsatellite marker, stability and polymorphism, obtain microsatellite marker.Its weak point is, is not suitable for detection and the application of the mark of japonicus family.
Summary of the invention
The object of the invention is to solve that the existing mark identifying different japonicus family is complicated, the defect of length consuming time and one conveniently japonicus molecule marking method is provided.
Another object of the present invention is the application providing a kind of japonicus molecule marker.
To achieve these goals, the present invention is by the following technical solutions:
A kind of japonicus molecule marking method, described japonicus molecule marking method comprises the following steps:
1) extract DNA genome: get japonicus and organize 150-200mg, extract DNA genome according to phenol chloroform extraction method after shredding, save backup at 0-4 DEG C;
2) pcr amplification: with step 1) the DNA genome that obtains is that template carries out pcr amplification, for special primer called after L10985 and the H11563 increased, obtains pcr amplification product; Wherein, L10985:5 '-TGAGGAGGTTTCGCAGTA-3 ';
H11563:5’-AGATGAGGGTGAGTGGGT-3’；
3) Product Identification: by step 2) pcr amplification product that obtains is separated at 1.5% agarose gel electrophoresis, and ethidium bromide staining, observes and Taking Pictures recording under gel imaging system.
In the technical program, molecule marker is with macromole biological between individuality, the genetic marker based on the hereditary material DNA nucleotide sequence variation of especially organism.Its development creates revolutionary impact to zoogenetics, and with traditional genetic marker if morphological markers, cytological marker are compared with isoenzyme mark, whole genomic genovariation can directly be observed and be probed into molecular marking technique fast.Have codominance in addition, quantity is extremely many, and polymorphism is high, not by advantages such as developmental stage, season, environmental restraint.
In assistant breeding process, utilize DNA molecular marker technology can carry out assisted Selection, shortening the breeding cycle to the important economical trait of japonicus (comprising disease-resistant, degeneration-resistant and breeding high-yield kind) closely linked gene, greatly raise the efficiency.
As preferably, step 2) in PCR reaction system cumulative volume be 50 μ L, comprise DNA genome 50-100mg, 10 × Buffer 5-10 μ L, dNTP 0.2-0.3mmol/L, each 0.2-0.4 of special primer L10985, H11563 μm ol/L, Taq plus archaeal dna polymerase 2-4U and magnesium chloride 1.5-1.8mmol/L.
As preferably, PCR reaction conditions is: 94 DEG C of denaturation 4min, 94 DEG C of sex change 30s, 50 DEG C of annealing 45s, 72 DEG C extend 45s, after 30 circulations, extend 7min at 72 DEG C.
As preferably, pcr amplification product is separated at 1.5% agarose gel electrophoresis, and electrophoretic buffer is 0.5 × TBE, its pH is 8.0, electrophoresis time is 1-1.5h, adds ethidium bromide and dyes to final concentration 500ng/mL, then observes and Taking Pictures recording under gel imaging system.
An application for japonicus molecule marker, described in be applied as the analysis of screening japonicus Germplasm Resources Diversity, analysis of genetic diversity, Parentage determination, molecular population genetics research, genetic map construction, important economical trait location, functional gene research and assist japonicus molecular genetic breeding or cultivation.
In the technical program, molecular marking technique can be found and detect and the gene of disease-related on DNA level, in assistant breeding process, utilize DNA molecular marker technology can carry out assisted Selection to the important economical trait of japonicus (comprising disease-resistant, degeneration-resistant and breeding high-yield kind) closely linked gene, shortening the breeding cycle, greatly raise the efficiency.Thus detection is made to some disease, and judge.Molecular marking technique also can detect pathogenic bacteria, classify and the qualification of genetic affinity, grasps the survival condition of pathogenic bacterium different times, thus effectively controls and avoid the outburst of the extensive disease of japonicus.
Concerning genetic diversity and phylogeny research, the precondition of cultivating good character kind is abundant genetic diversity resource, and abundant genetic diversity resource also can bring huge economic benefit for aquaculture, application molecular marking technique, can effectively analyze in japonicus kind or middle phylogeny and genetic diversity.
DNA molecular marker can make the japonicus genetic linkage maps of high-density and high coverage rate constantly perfect, the field such as the assignment of genes gene mapping, genetic breeding is significant, and the mark of important economical trait gene and clone, excellent to the heredity of cultivation japonicus, cultivate the healthy kind of good quality and high output and have a very big significance.
Beneficial effect of the present invention:
1) the present invention can obtain the heritable variation polymorphism collection of illustrative plates of the genetic marker locus of japonicus fast, and method is simple and efficient, can observe result intuitively;
2) the present invention is mainly used in the structure of genetic marker, genealogical identification and genetic map between japonicus colony;
3) the present invention detects rapidly, expense is low, and purposes is more extensive.
Embodiment
Below in conjunction with specific embodiment, the present invention will be further explained:
Embodiment 1
A kind of japonicus molecule marking method, described japonicus molecule marking method comprises the following steps:
1) DNA genome is extracted: get japonicus and organize 150mg, after shredding, put into 1.5mL centrifuge tube, add 500 μ l damping fluid (10mmol/L Tris-HCl, pH 8.0; 50mmol/L EDTA, pH 8.0), adding final concentration after mixing is the SDS of 1% and the Proteinase K of 200 μ g/ml, 55 DEG C of water-bath 3h; The centrifugal 5min of 12000 turns/min, gets supernatant; The saturated phenol of isopyknic Tris-with supernatant liquor is added: chloroform: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, the saturated phenol of Tris-: chloroform: primary isoamyl alcohol volume ratio is 25:24:1, then the centrifugal 5min of 12000 turns/min, get supernatant liquor, isopyknic chloroform is added with supernatant liquor: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, chloroform: the volume ratio of primary isoamyl alcohol is 24:1, and the centrifugal 5min of 5000 turns/min, gets supernatant liquor; In supernatant liquor, add the dehydrated alcohol precipitation of twice supernatant volume, then 70% washing with alcohol precipitation twice, dissolves with TBE; OD260, OD280 value of ultraviolet spectrophotometer working sample DNA, determines its concentration and purity, and 4 DEG C save backup; Totally 17 samples;
2) pcr amplification: with step 1) the DNA genome that obtains is that template carries out pcr amplification, for special primer called after L10985 and the H11563 increased, obtains pcr amplification product; Wherein, L10985:5 '-TGAGGAGGTTTCGCAGTA-3 ';
H11563:5 '-AGATGAGGGTGAGTGGGT-3 '; Wherein, PCR reaction system cumulative volume is 50 μ L, comprises DNA genome 100mg, 10 × Buffer 10 μ L, dNTP 0.3mmol/L, each 0.4 μm of ol/L, Taq plus archaeal dna polymerase 4U and magnesium chloride 1.8mmol/L of special primer L10985, H11563; PCR reaction conditions is: 94 DEG C of denaturation 4min, 94 DEG C of sex change 30s, 50 DEG C of annealing 45s, 72 DEG C of extension 45s, after 30 circulations, extend 7min at 72 DEG C;
3) Product Identification: by step 2) pcr amplification product that obtains is separated at 1.5% agarose gel electrophoresis, electrophoretic buffer is 0.5 × TBE, its pH is 8.0, electrophoresis time is 1.5h, add ethidium bromide to dye to final concentration 500ng/mL, then observe and Taking Pictures recording under gel imaging system.
Embodiment 2
A kind of japonicus molecule marking method, described japonicus molecule marking method comprises the following steps:
1) DNA genome is extracted: get japonicus and organize 180mg, after shredding, put into 1.5mL centrifuge tube, add 500 μ l damping fluid (10mmol/L Tris-HCl, pH 8.0; 50mmol/L EDTA, pH 8.0), adding final concentration after mixing is the SDS of 1% and the Proteinase K of 200 μ g/ml, 55 DEG C of water-bath 3h; The centrifugal 5min of 12000 turns/min, gets supernatant; The saturated phenol of isopyknic Tris-with supernatant liquor is added: chloroform: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, the saturated phenol of Tris-: chloroform: primary isoamyl alcohol volume ratio is 25:24:1, then the centrifugal 5min of 12000 turns/min, get supernatant liquor, isopyknic chloroform is added with supernatant liquor: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, chloroform: the volume ratio of primary isoamyl alcohol is 24:1, and the centrifugal 5min of 5000 turns/min, gets supernatant liquor; In supernatant liquor, add the dehydrated alcohol precipitation of twice supernatant volume, then 70% washing with alcohol precipitation twice, dissolves with TBE; OD260, OD280 value of ultraviolet spectrophotometer working sample DNA, determines its concentration and purity, and 4 DEG C save backup; Totally 17 samples;
2) pcr amplification: with step 1) the DNA genome that obtains is that template carries out pcr amplification, for special primer called after L10985 and the H11563 increased, obtains pcr amplification product; Wherein, L10985:5 '-TGAGGAGGTTTCGCAGTA-3 ';
H11563:5 '-AGATGAGGGTGAGTGGGT-3 '; Wherein, PCR reaction system cumulative volume is 50 μ L, comprises DNA genome 50mg, 10 × Buffer 5 μ L, dNTP 0.2mmol/L, each 0.2 μm of ol/L, Taq plus archaeal dna polymerase 2U and magnesium chloride 1.5mmol/L of special primer L10985, H11563; PCR reaction conditions is: 94 DEG C of denaturation 4min, 94 DEG C of sex change 30s, 50 DEG C of annealing 45s, 72 DEG C of extension 45s, after 30 circulations, extend 7min at 72 DEG C;
3) Product Identification: by step 2) pcr amplification product that obtains is separated at 1.5% agarose gel electrophoresis, electrophoretic buffer is 0.5 × TBE, its pH is 8.0, electrophoresis time is 1.2h, add ethidium bromide to dye to final concentration 500ng/mL, then observe and Taking Pictures recording under gel imaging system.
Embodiment 3
A kind of japonicus molecule marking method, described japonicus molecule marking method comprises the following steps:
1) DNA genome is extracted: get japonicus and organize 200mg, after shredding, put into 1.5mL centrifuge tube, add 500 μ l damping fluid (10mmol/L Tris-HCl, pH 8.0; 50mmol/L EDTA, pH 8.0), adding final concentration after mixing is the SDS of 1% and the Proteinase K of 200 μ g/ml, 55 DEG C of water-bath 3h; The centrifugal 5min of 12000 turns/min, gets supernatant; The saturated phenol of isopyknic Tris-with supernatant liquor is added: chloroform: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, the saturated phenol of Tris-: chloroform: primary isoamyl alcohol volume ratio is 25:24:1, then the centrifugal 5min of 12000 turns/min, get supernatant liquor, isopyknic chloroform is added with supernatant liquor: the extracting of primary isoamyl alcohol mixed solution once in supernatant liquor, chloroform: the volume ratio of primary isoamyl alcohol is 24:1, and the centrifugal 5min of 5000 turns/min, gets supernatant liquor; In supernatant liquor, add the dehydrated alcohol precipitation of twice supernatant volume, then 70% washing with alcohol precipitation twice, dissolves with TBE; OD260, OD280 value of ultraviolet spectrophotometer working sample DNA, determines its concentration and purity, and 4 DEG C save backup; Totally 17 samples;
2) pcr amplification: with step 1) the DNA genome that obtains is that template carries out pcr amplification, for special primer called after L10985 and the H11563 increased, obtains pcr amplification product; Wherein, L10985:5 '-TGAGGAGGTTTCGCAGTA-3 ';
H11563:5 '-AGATGAGGGTGAGTGGGT-3 '; Wherein, PCR reaction system cumulative volume is 50 μ L, comprises DNA genome 80mg, 10 × Buffer 8 μ L, dNTP 0.3mmol/L, special primer L10985, H11563 each 0.3 μm of ol/L, Taq plus archaeal dna polymerase 3U, magnesium chloride 1.6mmol/L; PCR reaction conditions is: 94 DEG C of denaturation 4min, 94 DEG C sex change 30s and 50 DEG C annealing 45s, 72 DEG C of extension 45s, after 30 circulations, extend 7min at 72 DEG C;
3) Product Identification: by step 2) pcr amplification product that obtains is separated at 1.5% agarose gel electrophoresis, electrophoretic buffer is 0.5 × TBE, its pH is 8.0, electrophoresis time is 1h, add ethidium bromide to dye to final concentration 500ng/mL, then observe and Taking Pictures recording under gel imaging system.
The PCR primer of embodiment 1-3 is sent to Shanghai Sheng Gong biotechnology company limited, checks order on sequenator at ABI 3730 after purifying.The sequence obtained by confirming the gene fragment of gained after BLAST Homology search in NCBI, and downloads the analysis of cytb sequence for phylogenetic relationship of other prawn species after manual synchronizing from GenBank database.Utilize the variant sites of MEGA3.1 software analysis DNA sequence dna and aminoacid sequence, calculate based composition ratio and genetic distance and constructing system evolutionary tree; DnaSP4.0 software is utilized to carry out the calculating of polymorphism index, Population Differentiation index and gene flow; Utilize DAMBE computed in software nucleotide diversity index.
Result is as follows:
The genetic diversity parameter of japonicus 4 colonies is as shown in table 1, Fujian colony has the highest level of genetic diversity, its haplotype diversity and nucleotide diversity are respectively 1.0 and 0.0137, its haplotype diversity of Zhejiang colony that level of genetic diversity is minimum and nucleotide diversity are respectively 0.874 and 0.0045, comprehensive parameters is known, and japonicus 4 geographical population level of genetic diversity are Fujian colony, Guangdong colony, Taiwan population and Zhejiang colony from high to low respectively.Check order row after manual synchronizing, find 52 variant sites altogether, account for 9.42% of site of analysis sum, the nucleotide diversity index of each variant sites is 0.014-0.198, comprises 38 brief informative sites in variant sites.These variant sites comprise 39 conversion sites and 14 transversion sites (table 2), do not insert and deletion segment, and wherein the conversion of institute of the Fujian colony analytical sequence that diversity level is the highest and transversion number reach 30 and 12 respectively.
Utilize the synonym substitution rate (dS) in MEGA computed in software japonicus cytb gene and synonymous substitution rate (dN), the dN/dS of full sequence is 0.130 (table 3).Adopt the Z test of single tail to carry out negative select (dN<dS) to check, replace due to synonym and replace apparently higher than non-synonym, check the P<0.01 obtained, reach pole conspicuous level, illustrate japonicus during evolution mtDNA-cytb gene be subject to the negative impact selected.
The genetic diversity parameter of table 1 japonicus 4 colonies
Utilize the genetic distance between MEGA computed in software colony and added up the variant sites number between colony, therefrom known, genetic distance between 4 colonies is between 0.004-0.012, wherein Fujian colony and Guangdong population genetic distance are 0.012 to the maximum, the genetic distance of Taiwan population and Zhejiang colony is minimum is 0.005, and the average genetic between all individualities is 0.009.And the genetic distance calculated in each colony between individuality, the intragroup genetic distance in Guangdong, Taiwan, Fujian and Zhejiang is respectively 0.008,0.005,0.014 and 0.005, and this has similar conclusion with the level of genetic diversity of table 1 Zhong Ge colony.Between the colony listed in table 2, variant sites digital display shows to there is more variant sites between Fujian colony and other colonies, there is larger genetic variation and genetic differentiation.
As can be seen from the genetic differentiation coefficient between 4 colonies calculated and gene flow data (table 3), slight genetic variation and genetic differentiation (Fst<0.05) is there is between Taiwan population and Zhejiang colony, Taiwan population and Fujian colony, between Zhejiang colony and Fujian colony, there is the genetic variation and genetic differentiation (Fst>0.15) of height, between other colonies, all there is the genetic variation and genetic differentiation (0.05<Fst<0.15) of moderate.Average genetic differentiation coefficient is 0.1004, and show that the genetic variation and genetic differentiation of about 10% comes between colony, the genetic variation and genetic differentiation of about 90% comes from colony.Gene flow (N
m) show that more greatly genetic variation and genetic differentiation degree is lower, the therefore same genetic variation and genetic differentiation level illustrated between each colony.
Relative Hereditary distance (under diagonal lines) and variant sites number (under diagonal lines, total variant sites/total variant sites) between table 2 japonicus 4 colony
| Colony stocks | Guangdong | Taiwan | Fujian | Zhejiang |
| Guangdong | 4/22 | 11/44 | 6/28 |
| Taiwan | 0.007 | 10/45 | 7/27 | |
| Fujian | 0.012 | 0.011 | 15/48 | |
| Zhejiang | 0.007 | 0.005 | 0.011 |
Population genetic coefficient of differentiation Fst (under diagonal lines) between table 3 japonicus 4 colony and gene flow Nm (on diagonal lines)
| Colony stocks | Guangdong | Taiwan | Fujian | Zhejiang |
| Guangdong | 2.90 | 1.93 | 4.55 | |
| Taiwan | 0.0794 | 1.11 | 13.15 | |
| Fujian | 0.1143 | 0.1833 | 1.37 | |
| Zhejiang | 0.0524 | 0.0185 | 0.1545 |
With reference to the level of genetic diversity of each colony that molecule marker obtains, the hybrid experiment of 8 combinations has been carried out in the basic population design utilizing 4 japonicus to breed, repeatedly mass mating experiment has been carried out from March, 2007 in June, 2007, finally obtain the success of japonicus colony hybridization, obtain hybridization shrimp seedling 8,650,000 tail in May, 2007.And carry out the extensive hybrid experiment of second time in May, 2007, simultaneously and travel the comparison test of 8 cross combinations.
On June 28th, 2007 puts various combination hybridization prawn seedling in a suitable place to breed in Zhoushan Lv Yuan aquaculture company limited (Dinghai is long white) and contrasts original seed prawn seedling in 8 test tanks, test area 250 × 8m
2, breeding density 1.6 ten thousand tails/pond.On September 12nd, 2007, basic condition was in table 4 to 8 pond harvestings and sampling Detection 50 tails/pond.Wherein 7# pond (G ♀ × T ♂) hybridizes japonicus average physical total length 14.6cm, mean body weight 12.4g, average yield per mu 354.3kg; (T ♀ × G ♂) hybridizes japonicus average physical total length 16.2cm in 8# pond, mean body weight 14.2g, average yield per mu 374.1kg, be all better than other combination through contrast discovery 7#, 8# pond cross combination speed of growth, unit output, shrimp is well-proportioned, size is even.
Table 4 Different Cross Combinations culture experiment basic condition table
Note: G Guangdong, F Fujian, catwalk gulf.
SEQUENCE LISTING
<110> Zhejiang Marine Development Research Institute
<120> japonicus molecule marking method and application
<130> ZH10013
<160> 2
<170> PatentIn version 3.3
<210> 1
<211> 18
<212> DNA
<213> synthetic
<400> 1
tgaggaggtt tcgcagta 18
<210> 2
<211> 18
<212> DNA
<213> synthetic
<400> 2
agatgagggt gagtgggt 18
Claims (5)
1. a japonicus molecule marking method, is characterized in that, described japonicus molecule marking method comprises the following steps:
1) extract DNA genome: get japonicus and organize 150-200mg, extract DNA genome according to phenol chloroform extraction method after shredding, save backup at 0-4 DEG C;
2) pcr amplification: with step 1) the DNA genome that obtains is that template carries out pcr amplification, for special primer called after L10985 and the H11563 increased, obtains pcr amplification product; Wherein, L10985:5 '-TGAGGAGGTTTCGCAGTA-3 ';
H11563:5’-AGATGAGGGTGAGTGGGT-3’；
3) Product Identification: by step 2) pcr amplification product that obtains is separated at 1.5% agarose gel electrophoresis, and ethidium bromide staining, observes and Taking Pictures recording under gel imaging system.
2. a kind of japonicus molecule marking method according to claim 1, it is characterized in that, step 2) in PCR reaction system cumulative volume be 50 μ L, comprise DNA genome 50-100mg, 10 × Buffer 5-10 μ L, dNTP 0.2-0.3mmol/L, each 0.2-0.4 of special primer L10985, H11563 μm ol/L, Taq plus archaeal dna polymerase 2-4U and magnesium chloride 1.5-1.8mmol/L.
3. a kind of japonicus molecule marking method according to claim 1 and 2, it is characterized in that, PCR reaction conditions is: 94 DEG C of denaturation 4min, 94 DEG C of sex change 30s, 50 DEG C of annealing 45s, 72 DEG C of extension 45s, after 30 circulations, extend 7min at 72 DEG C.
4. a kind of japonicus molecule marking method according to claim 1, it is characterized in that, pcr amplification product is separated at 1.5% agarose gel electrophoresis, electrophoretic buffer is 0.5 × TBE, its pH is 8.0, electrophoresis time is 1-1.5h, adds ethidium bromide and dyes to final concentration 500ng/mL, then observes and Taking Pictures recording under gel imaging system.
5. the application of an a kind of japonicus molecule marker as claimed in claim 1, it is characterized in that, described in be applied as the analysis of screening japonicus Germplasm Resources Diversity, analysis of genetic diversity, Parentage determination, molecular population genetics research, genetic map construction, important economical trait location, functional gene research and assist japonicus molecular genetic breeding or cultivation.
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| CN113584187A (en) * | 2021-09-10 | 2021-11-02 | 中国水产科学研究院黄海水产研究所 | Molecular marker A2629 for screening penaeus japonicus with low temperature resistance, amplification primer and application thereof |
| CN113584188A (en) * | 2021-09-10 | 2021-11-02 | 中国水产科学研究院黄海水产研究所 | Low-temperature-resistant molecular marker C6101 of penaeus japonicus and application |
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| CN113621714A (en) * | 2021-09-10 | 2021-11-09 | 中国水产科学研究院黄海水产研究所 | Low-temperature-resistant molecular marker A257 of penaeus japonicus and application thereof |
| CN113736891A (en) * | 2021-09-10 | 2021-12-03 | 中国水产科学研究院黄海水产研究所 | Molecular marker G2997 for rapidly identifying low-temperature tolerant variety of penaeus japonicus and application thereof |
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| CN108070663A (en) * | 2017-12-06 | 2018-05-25 | 中国水产科学研究院南海水产研究所 | Differentiate the molecular labeling primer and method of the penaeus penicillatus young and During Larvae Rearing of Penaeus Merguiensis De Man |
| CN108467892A (en) * | 2018-01-05 | 2018-08-31 | 广西壮族自治区海洋研究所 | A kind of screening technique of 2 kinds of morphological variation body candidate sequences of Marsupenaeus japonicus |
| CN113584187A (en) * | 2021-09-10 | 2021-11-02 | 中国水产科学研究院黄海水产研究所 | Molecular marker A2629 for screening penaeus japonicus with low temperature resistance, amplification primer and application thereof |
| CN113584188A (en) * | 2021-09-10 | 2021-11-02 | 中国水产科学研究院黄海水产研究所 | Low-temperature-resistant molecular marker C6101 of penaeus japonicus and application |
| CN113604587A (en) * | 2021-09-10 | 2021-11-05 | 中国水产科学研究院黄海水产研究所 | Molecular marker T5198 for rapidly identifying low-temperature tolerant variety of penaeus japonicus and application thereof |
| CN113621714A (en) * | 2021-09-10 | 2021-11-09 | 中国水产科学研究院黄海水产研究所 | Low-temperature-resistant molecular marker A257 of penaeus japonicus and application thereof |
| CN113736891A (en) * | 2021-09-10 | 2021-12-03 | 中国水产科学研究院黄海水产研究所 | Molecular marker G2997 for rapidly identifying low-temperature tolerant variety of penaeus japonicus and application thereof |
| CN113736891B (en) * | 2021-09-10 | 2022-04-22 | 中国水产科学研究院黄海水产研究所 | Molecular marker G2997 for rapidly identifying low-temperature tolerant variety of penaeus japonicus and application thereof |
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