CN106701948B - Method for identifying female parent sources of purple scallops, bay scallops and hybrid offspring - Google Patents
Method for identifying female parent sources of purple scallops, bay scallops and hybrid offspring Download PDFInfo
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Abstract
The invention relates to a method for identifying female parent sources of scallop adductor scallop, bay scallop and filial generation, which is a marked primer mtDNA-Z designed based on the difference of mitochondrial DNA sequences, and is characterized in that the source of the female parent source is respectively expressed by the following primer sequences mtDNA-Z-F at the left end: 5'-TATGAGGTGTCCCCCAAGTC-3' and right primer sequence mtDNA-Z-R: 5'-ACTGGCAGACAAACAAATCGT-3' are added to the mixture to form a mixture, the extracted DNA of the purple scallop, the bay scallop and the hybrid scallop is taken as a template, and the primer is utilized to carry out PCR amplification on the extracted DNA, if a PCR specific band appears at a position of near 460bp, determining that the source of the female parent mitochondrial gene is purple scallop, if no PCR specific band appears at the position of near 460bp, the source of the female parent mitochondrial gene is determined to be bay scallop, the invention eliminates the influence of nuclear genome DNA, does not need to separately extract the mitochondrial DNA for identification, therefore, the parent sources of the purple scallops, bay scallops and hybrid offspring can be identified quickly and efficiently, the method is simple and convenient, the cycle time is short, can be carried out in large batch, and provides technical support and guarantee for ensuring the smooth progress of the breeding process and confirming the pedigree of the offspring.
Description
Technical Field
The invention belongs to a descendant identification technology of scallop crossbreeding, and particularly relates to an identification method of female parent sources of purple scallops, bay scallops and hybrid descendants.
Background
Argopecten irradians (Argopecten irradians) has been introduced from the United states in 1982, the yield of the Argopecten irradians is increased year by year, and the Argopecten irradians is one of the most important cultured shellfish in China, but the germplasm degradation problem appearing in recent years becomes a serious obstacle for restricting the development of scallop culture industry in China.
Purple scallops (Argopecten purpuratus) are excellent economic shellfish on the coast of the Pacific ocean in south America, have medium sizes, are widely cultivated in Chilean and Peru, and make Chilean a second-generation great country for scallop cultivation. In order to improve the germplasm of bay scallops and improve the economic yield of the scallop breeding industry, Wangchunde is equal to the first filial generation which is successfully hybridized with bay scallops and is introduced from Peru in 2008, and two filial generations with extremely obvious growth advantages are cultured, namely the purple-sea hybrid scallop (purple scallop ovum and bay scallop sperm) and the sea-purple hybrid scallop (bay scallop ovum and scallop sperm) (Wangchund et al, 2009; Wang et al, 2011). However, the purple scallops and bay scallops are both hermaphrodite animals, and both the scallops discharge sperms and eggs at the same time during breeding, so that the problem of pollution of the sperms or the eggs exists in breeding practice, and in order to ensure smooth progress of breeding and confirm the pedigrees of offspring, whether a certain offspring is a real hybrid offspring or not and the source of the male parent and the female parent of the offspring is often required to be determined, so that a method is required to be established for rapidly identifying the source of the parents, particularly the source of the female parent.
Although the identification method using molecular markers (such as microsatellite markers) from a nuclear genome can also be used for identifying whether filial generations are true hybrids, the source of the parents cannot be distinguished, so that the filial generations are disordered in pedigree, the excellent lines are difficult to screen, and the like.
Disclosure of Invention
The invention aims to provide a method for identifying the source of female parent of purple scallop, bay scallop and hybrid progeny, which is a marked primer designed based on the difference of mitochondrial DNA sequences and is used for identifying the source of female parent of purple scallop, bay scallop and hybrid progeny so as to make up the defects of the prior art.
Research has found that mitochondrial genomes strictly follow maternal inheritance rules and lack gene recombination, mitochondria of filial generations must come from their parents, and thus the maternal origin of a hybrid can be determined by identifying the source of its mitochondrial DNA. According to the difference characteristics of mitochondrial genomes of bay scallops and purple scallops, the invention specially designs a pair of specific primers so as to identify the type of mitochondrial genes of hybrid offspring by a PCR method, thereby realizing the rapid identification of female parent sources of hybrid scallops.
The invention is based on the following idea: the existing research shows that the mitochondrial genome of the scallop follows strict maternal inheritance, namely the mitochondrial gene of the offspring is completely the same as the mitochondrial genotype of the female parent and is unrelated to the mitochondrial genotype of the male parent; particularly, the bay scallop and the purple scallop are all related species of Argopecten, and although the similarity of mitochondrial genome sequences is as high as 82.34 percent, specific primers can be designed according to a little difference of the mitochondrial gene sequences of the bay scallop and the purple scallop, so that the sources of filial generation mitochondrial genes can be distinguished, and technical support or guarantee is provided for smooth breeding work.
The invention is realized by the following technical scheme: the method for identifying the female parent sources of the purple scallops, bay scallops and hybrid offspring comprises the steps that the used marking primer is mtDNA-Z, and the marking primer is composed of a left end primer sequence mtDNA-Z-F: 5'-TATGAGGTGTCCCCCAAGTC-3' and right primer sequence mtDNA-Z-R: 5'-ACTGGCAGACAAACAAATCGT-3'.
The method for identifying the female parent sources of the purple scallops, bay scallops and hybrid offspring of the purple scallops and the bay scallops by using the labeled primers comprises the following steps:
(1) taking the adductor muscles of the purple scallops to be identified, bay scallops, hybrid offspring purple sea hybrid scallops and sea purple hybrid scallops respectively, directly extracting the whole genome DNA of the scallops by a conventional method, diluting the whole genome DNA to a final concentration of 100 ng/mu L by TE buffer solution, and storing the diluted whole genome DNA at-20 ℃ for later use;
(2) the extracted DNAs of the above scallops were used as templates, respectively, and PCR amplification was carried out using the above primer mtDNA-Z in a reaction system of 10. mu.L including 0.25U of Taq DNase (Takara Inc., Shiga, Japan) and 1 XPCR buffer (containing 1.5mM MgCl. of 1.5 mM)2) And 0.2mM dNTP mix, 1. mu.M each of upstream and downstream primers, 50ng of DNA template; the reaction procedure is as follows: pre-denaturation at 94 deg.C for 3min, pre-denaturation at 94 deg.C for 1min, pre-denaturation at 56 deg.C for 1min, pre-denaturation at 72 deg.C for 1min, and pre-denaturation for 30 cycles, and pre-extension at 72 deg.C for 10 min; performing 1% agarose gel electrophoresis on the PCR product, performing EB (electron beam) dyeing, and performing photographic observation by using a gel imaging system; for the purple scallops and bay scallops, if a PCR specific band appears at a position close to 460bp, the scallops are proved to be purple scallops, and if no PCR specific band appears at a position close to 460bp, the scallops are proved to be bay scallops; for purple sea hybrid scallops and sea purple hybrid scallops, if PCR specific bands appear at a position close to 460bp, the female parent is determined to be purple scallop, and if no PCR specific bands appear at a position close to 460bp, the female parent is determined to be bay scallop.
The invention has the following advantages:
(1) the primer design based on the mitochondrial DNA sequence of the invention eliminates the influence of nuclear genome DNA, so that the mitochondrial DNA does not need to be extracted independently for identification, a small amount of mitochondrial DNA in the whole genome extracted conventionally is utilized, the labeled primer mtDNA-Z can be used for PCR amplification, and an identification result can be obtained after agarose electrophoresis detection, so that the cycle time is short, the method is simple and convenient, and the method can be carried out in large batch, quickly and efficiently.
(2) The specific primer designed based on the mitochondrial DNA sequence can quickly, simply and conveniently identify two filial generations and two parents thereof.
Drawings
FIG. 1 is an electrophoresis diagram of products of Argopecten irradians and Argopecten irradians by the labeled primer mtDNA-Z PCR in example 1 of the present invention, wherein 1-6: argopecten irradians, 7-12 Argopecten irradians, and M DL 2000.
FIG. 2 is an electrophoresis diagram of hybrid generation purple scallop and purple scallop products obtained by PCR amplification of a labeled primer mtDNA-Z used in example 2 of the invention, wherein the ratio of 1-8: purple scallop, 9-16: argopecten irradians, M, Marker.
Detailed Description
Example 1 the results of the identification were performed using purple scallops and bay scallops as examples.
(1) The adductor muscles of 6 purple scallops and 6 bay scallops are respectively taken, the whole genome DNA is directly extracted by a Tiangen DNA extraction kit (Tiangen Biochemical technology Co., Ltd., Beijing), diluted by TE buffer solution to the final concentration of 100 ng/microliter and stored at the temperature of minus 20 ℃ for later use, and then the purity and the quality of the extracted whole genome DNA are ensured to meet the requirements through 1.2 percent agarose gel electrophoresis, EB staining and photographic observation of a gel imaging system.
(2) The DNA of the above-extracted purple scallop and Argopecten irradians was used as a template, and the above-mentioned primer mtDNA-Z was used to perform PCR amplification in a reaction system of 10. mu.L including 0.25U of Taq DNase (Takara Inc., Shiga, Japan) containing 1.5mM MgCl 21 XPCR buffer,0.2mM dNTP mix, 1. mu.M each of the upstream and downstream primers, 50ng of DNA template. The reaction procedure is as follows: pre-denaturation at 94 ℃ for 3min, at 94 ℃ for 1min, at 56 ℃ for 1min, at 72 ℃ for 1min, and 30 cycles; extension at 72 ℃ for 10 min. The PCR product was electrophoresed through 1% agarose gel, stained with EB, and the electrophoresis result was observed by a gel imaging system.
(3) The electrophoresis result shows that no band is amplified in all bay scallops (see figure 1), while a clear band is amplified in all purple scallops around 460bp, and the size of the band is consistent with the expected 463 bp. The amplified product in the purple scallop is purified, the sequencing result size is 460bp, DNAMAN comparison is carried out on the amplified product and a mitochondrial DNA sequence of the purple scallop, and the fragment similarity of the sequence and the mitochondrial DNA of the purple scallop is 97.81 percent (see table 1), so that the mtDNA-Z primer is prevented from being influenced by nuclear genome, and the amplified product is determined to be PCR amplification reaction carried out by taking the mitochondrial DNA of the purple scallop as a template. Obviously, the primers designed by the invention can identify the purple scallops and bay scallops.
TABLE 1 sequencing results comparison Table
Example 2 results of identification using purple-sea hybrid scallops and sea-purple hybrid scallops as examples.
(1) Taking 8 scallop adductor muscles of purple sea hybrid scallops (purple scallop (egg) × bay scallops (sperms)) and sea purple hybrid scallops (bay scallops (egg) × purple scallops (sperms)) respectively, extracting whole genome DNA by a Tiangen DNA extraction kit (Tiangen Biochemical technology Co., Ltd., Beijing), diluting the whole genome DNA to a final concentration of 100 ng/microliter by TE buffer solution and storing the final concentration at-20 ℃ for later use, carrying out 1.2% agarose gel electrophoresis on the extracted whole genome DNA, carrying out EB staining, and carrying out photographic observation by a gel imaging system so as to ensure that the purity and the quality of the extracted whole genome DNA meet requirements.
(2) The extracted DNA of the purple sea hybrid scallop and the extracted DNA of the sea purple hybrid scallop are used as templates, the primer pair is used for PCR amplification, the reaction system is 10 mu L, and the reaction system comprises 0.25U Taq DNase (Takara Inc., Shiga, Japan) and 1 XPCR buffer (containing 1.5mM MgCl2) 0.2mM dNTP mix, 1. mu.M each of the upstream and downstream primers, 50ng of DNA template. The reaction procedure is as follows: pre-denaturation at 94 ℃ for 3min, at 94 ℃ for 1min, at 56 ℃ for 1min, at 72 ℃ for 1min, and 30 cycles; extension at 72 ℃ for 10 min. The PCR product was electrophoresed through 1% agarose gel, stained with EB, and the electrophoresis result was observed by a gel imaging system.
(3) The electrophoresis result identification shows that all the purple-sea hybrid scallops amplify obvious bands near 460bp, the size of the bands is consistent with the expected 463bp, and the bands are not amplified in all the purple-sea hybrid scallops (see figure 2), which indicates that the designed primer pair identifies the female parent source of the hybrid offspring of the purple scallops and bay scallops without errors.
SEQUENCE LISTING
<110> Qingdao agricultural university
<120> method for identifying female parent sources of purple scallops, bay scallops and hybrid offspring
<160>1
<210>1
<211>463
<212>DNA
<213> purple scallop (Argopecten purpuratus)
<220>
<221>misc_feature
<400>1
tatgaggtgt cccccaagtc ttgggttttt gggggaggtt ataataggga tagggatttg 60
tagaattttt ccacaaggct atattttttg ttttattcta ctattctttt tcggtggtgc 120
aagaataatg gtgctttaca ccagggtaat acacggaagg ttttcgactg ccctggttcc 180
tggggggtct ggaattacga agtgtagtta tcttggtctt ttccatgggg tgccgttaat 240
tattttgttt tttttacctg cgttcttgtc tttgcgggct cttcggagcg gtctttaaga 300
agtagggcta aagcccacgc actgttgaca gagtaggaga ggaggaagta gaaaaccaga 360
ttaaatgggt ggcgagagcg atgagattgt acaagacccg ggataaaggc taagagaagg 420
ggtctcccga tcgaattagg gtacgatttg tttgtctgcc agt 463
Claims (1)
1. A method for identifying the source of female parent of purple scallop, bay scallop and their hybrid offspring is characterized in that the used marker primer is mtDNA-Z, which is composed of left end primer sequence mtDNA-Z-F and right end primer sequence mtDNA-Z-R, wherein the left end primer sequence mtDNA-Z-F is 5'-TATGAGGTGTCCCCCAAGTC-3', and the right end primer sequence mtDNA-Z-R is 5'-ACTGGCAGACAAACAAATCGT-3';
the method for identifying the female parent sources of the purple scallops, bay scallops and hybrid offspring of the purple scallops and the bay scallops comprises the following specific steps:
(1) taking adductor muscles of purple scallops, bay scallops, hybrid scallops and hybrid scallops to be identified, directly extracting whole genome DNA by a conventional method, diluting the whole genome DNA by TE buffer solution to a final concentration of 100 ng/mu L, and storing the diluted whole genome DNA at-20 ℃ for later use; wherein the hybrid scallop is the offspring of Argopecten irradians ovum and Argopecten irradians sperm;
(2) respectively taking the extracted DNA of the purple scallop, the bay scallop and the hybrid scallop as templates, and carrying out PCR amplification on the extracted DNA by using the primer mtDNA-Z, wherein the reaction system is 10 mu L, comprises 0.25U Taq DNase and contains 1.5mM MgCl21 x PCRbuffer of (1 x PCRbuffer),0.2mM dNTP mix, 1 μ M each of left and right primers, 50ng DNA template; the reaction procedure is as follows: pre-denaturation at 94 deg.C for 3min, pre-denaturation at 94 deg.C for 1min, pre-denaturation at 56 deg.C for 1min, pre-denaturation at 72 deg.C for 1min, and pre-denaturation for 30 cycles, and pre-extension at 72 deg.C for 10 min; carrying out 1% agarose gel electrophoresis on the PCR product, EB dyeing, and observing an electrophoresis result by a gel imaging system; if a PCR specific band appears at the position of near 460bp, the female parent is determined to be purple scallop, and if no PCR specific band appears at the position of near 460bp, the female parent is determined to be bay scallop.
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| CN109430126B (en) * | 2018-11-06 | 2020-09-15 | 广东海洋大学 | Method for cultivating new hybrid scallop variety of purple cuttlefish |
| CN109207613B (en) * | 2018-11-28 | 2022-03-04 | 宁波大学 | Molecular marker for identifying female parent source of hybrid individual of mytilus coruscus and Mediterranean mussel |
| CN109825599B (en) * | 2018-11-28 | 2022-08-09 | 宁波大学 | Method for identifying individual genetic background of hybrid of mytilus coruscus and Mediterranean mussel |
| CN115807109A (en) * | 2022-12-20 | 2023-03-17 | 中国海洋大学三亚海洋研究院 | PCR technology-based scallop variety identification method and specific primers thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1322845A (en) * | 2000-05-16 | 2001-11-21 | 北海道渔业协同组合连合会 | Method for analyzing phylogenetic ancestry of scallop |
| CN103602738A (en) * | 2013-11-12 | 2014-02-26 | 辽宁大学 | Multiple PCR (Polymerase Chain Reaction) primer and method for quickly identifying variety of scallops |
-
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1322845A (en) * | 2000-05-16 | 2001-11-21 | 北海道渔业协同组合连合会 | Method for analyzing phylogenetic ancestry of scallop |
| CN103602738A (en) * | 2013-11-12 | 2014-02-26 | 辽宁大学 | Multiple PCR (Polymerase Chain Reaction) primer and method for quickly identifying variety of scallops |
Non-Patent Citations (4)
| Title |
|---|
| Argopecten irradians irradians mitochondrion，complete genome;Li,H.等;《Genbank Database》;20150824;Accession：KT161262 * |
| Argopecten purpuratus mitochondrion，complete genome;Li,H.等;《Genbank Database》;20150824;Accession：KT161260 * |
| 栉孔扇贝、虾夷扇贝及其杂交子代线粒体COI和Cytb基因遗传多样性分析;于涛等;《海洋科学》;20160331;第40卷(第3期);摘要，第1.1、1.2、1.3、1.5、2.1、2.4节、图4 * |
| 紫扇贝与海湾扇贝种间杂交的研究;王春德等;《海洋科学》;20091031;第33卷(第10期);摘要，第84页左栏第1段至右栏第1段，第1.2、3.2节 * |
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