CN103409420B - Y-chromosome specific molecular marker of Nile tilapia, and genetic sex determination method and supermale producing method both based on molecular marker - Google Patents
Y-chromosome specific molecular marker of Nile tilapia, and genetic sex determination method and supermale producing method both based on molecular marker Download PDFInfo
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Abstract
The invention discloses a Y-chromosome specific molecular marker of Nile tilapia, wherein the nucleotide sequence of the Y-chromosome specific molecular marker is as shown in SEQ ID No.4; the Y-chromosome specific molecular marker can be commonly used for a plurality of strains of Nile tilapia; a genetic sex determination method of Nile tilapia is established based on the molecular marker, so that normal female fish XX, normal male fish XY, converted female fish XY, converted female fish YY and supermale fish YY can be differentiated from each other economically, rapidly and accurately; large-scale production of supermale fish fry can be realized through mating of YY supermale fish and the normal female fish XX based on the genetic sex determination method; as a result, the yield of culture is improved remarkably, the culture cost is reduced and the economic benefit is enhanced; besides, a method of producing the YY supermale fish through mating of the supermale fish YY and the converted female fish YY is also established by using the Y-chromosome specific molecular marker.
Description
Technical field
The invention belongs to molecular breeding technology field, relate to tilapia sex specific molecular marker, based on the tilapia genetic sex identification method of this molecule marker and the method based on this molecule marker production supermale fish.
Background technology
Tilapia is also known as African crucian, originate in the African continent and the Atlantic coast salt-fresh water waters, the Middle East, belonging to Osteichthyes Perciformes Callichthyidae, is tropic fishes, has the advantages such as reproductivity is strong, growth is fast, feeding habits are wide, feed request is low, lower oxygen concentration resistance, adaptability and disease resistance are strong.In recent decades, along with tilapia is introduced in succession in countries in the world, its one of main cultured fishes kind that become international.At present, the tilapia breed variety of China has Mozambique tilapia, bolti, Oreochromis aureus etc., wherein mainly bolti.China is the first producing country of tilapia and export State.
Because tilapia is short for sexual cycle, breeding is fast, female slower than male growth, tilapia excessive multiplication in cultivation, obtains fast, individual large, the marketable fish that neat specification, output are high of growth, needs to carry out single male Tilipia production.The method obtaining single male Tilipia at present on producing mainly contains three kinds: one, sex-reversal method, and namely obtain full milter with male hormone process fry, the method is simple and easy to do, but there is the suspicion of hormone residues, does not meet the requirement of pollution-free healthy cultivation; Two, interspecific hybridization method, as bolti ♀ × Oreochromis aureus ♂, the method is pollution-free, marketable fish performance is good, but needs parent very pure, obtains male ratio and is generally only 90-95%, still have a considerable amount of raun, also there is crossing barrier to a certain degree, the breeding cycle is long, and numerous seedling amount is on the low side; Three, three series mating method, also known as Genetic male tilapia technology (Genetically Male Tilapia, GMT), combine by sex-reversal and hybridizing method, with former system (XX ♀, XY ♂), transform system (XY ♀, YY ♀) and Male homozygous system (YY ♂) is supporting produces hereditary complete male tilapia.Concrete grammar is as follows: undifferentiated for sex juvenile fish seedling is carried out feminized treatment, transforms raun (XY ♀) by filial generation Test Identification; Raun (XY ♀) and former system milter (XY ♂) mating will be transformed, by filial generation Test Identification YY supermale fish; By YY supermale fish with transform raun (XY ♀) mating carry out feminized treatment, transform raun by filial generation Test Identification YY; YY is transformed raun and the mating of YY supermale fish, obtain full YY supermale fish; By YY supermale fish and former system raun (XX ♀) mating, produce full milter.Above-mentioned three line for hybrid seed production program more complicated, required time is also longer, but once obtain YY supermale fish and YY conversion raun, just can obtain YY supermale fish (YY ♀ × YY ♂) in a large number, be a kind of method comparing science.But because prior art can only judge the genotype of parent after test cross experiment for sex ratio according to tester, complex operation is time-consuming, labour intensity is large, seriously hinder applying of GMT, therefore, the key of three series mating method is, how to distinguish normal raun XX ♀, normal milter XY ♂ quickly and efficiently, transforms raun XY ♀, transforms raun YY ♀ and supermale fish YY ♂.
Along with development and the maturation of molecular marking technique, judge with the closely linked molecule marker of target gene molecular marker assisted selection (the Marker-Assisted Selection whether target gene exists by analyzing, MAS) technology is identified for the genetic sex of tilapia, and its key is to filter out the special molecule marker of tilapia sex.By hero (female) property specific DNA molecule marker chain with sex chromosome (sex) in screening tilapia genome, greatly can simplify the Production Flow Chart of GMT by MAS technology, reduce production cost, increase the benefit.Contriver place seminar is in early-stage Study, filter out Y-chromosome specific molecular marker of Nile tilapia NtY1 and NtY2 and X chromosome specific molecular marker NtX1, and the genetic sex identification method (Chinese patent ZL201010501061.8) of bolti is established based on above-mentioned molecule marker, also develop another bolti sex specific molecular marker (in the present invention called after marker2) on this basis.But above-mentioned sex specific molecular marker is all not ideal enough, slightly far away with the distance in bolti sex determination site, be only applicable to the bolti strain of screening this mark, and the bolti of multiple strain can not be common to.
Summary of the invention
In view of this, an object of the present invention is to provide a kind of Y-chromosome specific molecular marker of Nile tilapia, can be common to the bolti of multiple strain; Two of object is to provide a kind of method of carrying out the qualification of bolti genetic sex based on above-mentioned molecule marker, can distinguish normal raun XX ♀, normal milter XY ♂ economical, quickly and accurately, transforms raun XY ♀, transform raun YY ♀ and supermale fish YY ♂; Three of object is to provide a kind of method based on above-mentioned molecule marker fast and efficient production supermale fish, to cultivate by YY supermale fish the large-scale production that route realizes full milter fry, significantly improves cultured output, reduces aquaculture cost, increase economic efficiency.
After deliberation, the invention provides following technical scheme:
1. Y-chromosome specific molecular marker of Nile tilapia, its nucleotide sequence is as shown in SEQ ID No.4.
2. the molecule marking method of bolti genetic sex qualification, comprises the following steps:
A.PCR increases: the fin ray getting bolti to be measured, extracts genomic dna; With gained genomic dna for template, adopt the upstream primer shown in SEQ ID No.1 and the downstream primer shown in SEQ ID No.2, pcr amplification bolti sex specific molecular marker: the nucleotide sequence of X chromosome specific molecular marker is as shown in SEQ ID No.3, and the nucleotide sequence of Y chromosome specific molecular marker is as shown in SEQ ID No.4;
B. result judges: from the genomic dna of bolti to be measured, only amplify X chromosome specific molecular marker, judge that this bolti to be measured is the raun in heredity, sex chromosome genotype is XX; From the genomic dna of bolti to be measured, only amplify Y chromosome specific molecular marker, judge that this bolti to be measured is the supermale fish in heredity, sex chromosome genotype is YY; From the genomic dna of bolti to be measured, amplify X chromosome specific molecular marker and Y chromosome specific molecular marker, judge that this bolti to be measured is the milter in heredity, sex chromosome genotype is XY simultaneously.
3. adopt the molecule marking method of bolti genetic sex qualification to produce the method for supermale fish, comprise the following steps:
A. in the post-coitum of bolti raun XX and milter XY generation, is carried out feminized treatment, adopt the molecule marking method of bolti genetic sex qualification, filter out the conversion raun that sex chromosome genotype is XY;
B. XY is transformed raun and the mating of bolti XY milter, adopt the molecule marking method of bolti genetic sex qualification, from offspring, filter out the supermale fish that sex chromosome genotype is YY; Separately get XY to transform raun and bolti XY milter post-coitum generation and carry out feminized treatment, the molecule marking method adopting bolti genetic sex to identify, filters out the conversion raun that sex chromosome genotype is YY;
C. YY supermale fish and YY are transformed raun mating, obtain full YY supermale fish;
D. YY supermale fish offspring is carried out feminized treatment, obtain YY and transform raun;
E. repeating step C and D, transforms raun mating by YY supermale fish and YY and produces YY supermale fish.
After employing aforesaid method ensures to continue to obtain YY supermale fish, by YY supermale fish and the mating of bolti XX raun, complete male marketable fish can be produced in a large number.
Beneficial effect of the present invention is: genetic linkage analysis shows, when getting rid of natural sex and reversing interference, bolti X of the present invention and Y chromosome specific molecular marker (in the present invention difference called after sdX and sdY) are 0cM with the genetic distance in bolti sex determination site, obviously be better than bolti X described in Chinese patent ZL201010501061.8 and Y chromosome specific molecular marker NtX1 and NtY2, and another bolti sex specific molecular marker marker2 that this seminar filters out, the bolti of multiple strain can be common to, as to from Hainan, 98 tail boltis of 4 different lines of four the tilapia nurseries in Guangxi and Guangdong carry out double blinding inspection, genetic sex and phenotypic sex identify that identical rate is 90% ~ 100%, based on the bolti genetic sex identification method that X of the present invention and Y chromosome specific molecular marker sdX/sdY builds, normal raun XX ♀, normal milter XY ♂ can be distinguished economical, quickly and accurately, transform raun XY ♀, transform raun YY ♀ and supermale fish YY ♂, the method of YY supermale fish is produced in the YY ♀ set up based on above-mentioned bolti genetic sex identification method and YY ♂ mating, can fast and efficient production YY supermale fish, and then the large-scale production of full milter fry is realized by YY supermale fish and normal raun XX ♀ mating, significantly improve cultured output, reduce aquaculture cost, increase economic efficiency.
Accompanying drawing explanation
In order to make object of the present invention, technical scheme and beneficial effect clearly, the invention provides following accompanying drawing and being described:
Fig. 1 is the sequence alignment figure of X of the present invention and Y chromosome specific molecular marker.
Fig. 2 is that application X of the present invention and Y chromosome specific molecular marker carry out the result of genetic sex qualification to 5 tail bolti raun XX, 5 tail bolti milter XY and 5 tail bolti supermale fish YY, and M represents DNA molecular amount standard DL2000.
Fig. 3 is that application X of the present invention and Y chromosome specific molecular marker carry out the result of genetic sex qualification to 19 tail XX ♀ × XX ♂ bolti filial generations, wherein 1-19 is XX ♀ × XX ♂ filial generation, XX, XY, YY are contrast, and M represents DNA molecular amount standard DL2000.
Fig. 4 is that application X of the present invention and Y chromosome specific molecular marker carry out the result of genetic sex qualification to 19 tail XX ♀ × YY ♂ bolti filial generations, wherein 1-19 is XX ♀ × YY ♂ filial generation, XX, XY, YY are contrast, and M represents DNA molecular amount standard DL2000.
Fig. 5 is that application X of the present invention and Y chromosome specific molecular marker carry out the result of genetic sex qualification to 19 tail YY ♀ × YY ♂ bolti filial generations, wherein 1-19 is YY ♀ × YY ♂ filial generation, XX, XY, YY are contrast, and M represents DNA molecular amount standard DL2000.
Fig. 6 is X of the present invention and Y chromosome specific molecular marker and the sex determination site linkage map on sex chromosome.
Embodiment
Below in conjunction with accompanying drawing, the preferred embodiments of the present invention are described in detail.The experimental technique of unreceipted actual conditions in preferred embodiment, usually conveniently condition, such as the Molecular Cloning: A Laboratory guide (third edition, J. the work such as Pehanorm Brooker, Huang Peitang etc. translate, Science Press, 2002) described in condition, or according to the condition that manufacturer advises.
One, the acquisition of bolti sex specific molecular marker
1, the extraction of genomic dna
Clip bolti (is the Japanese strain bolti that this laboratory is introduced, its female parent draws from Egyptian, male parent is drawn from Malaysia) fin ray 10mg, put homogenate in lysate [10mM Tris-HCl (pH8.0)+100mM EDTA (pH8.0)+100mM NaCl+5mg/ml SDS] 600 μ l, add the ribonuclease A (RnaseA) that Proteinase K that final concentration is 20mg/ml and final concentration are 100 μ g/ml, 55 DEG C of water-bath digestion are to clarification, add phenol chloroform-primary isoamyl alcohol (volume ratio is 25:24:1) mixed solution 600 μ l again, after abundant mixing in 12000rpm centrifugal 10 minutes, Aspirate supernatant, again add phenol chloroform-primary isoamyl alcohol (volume ratio is 25:24:1) mixed solution extracting DNA, centrifugal, Aspirate supernatant, add ice ethanol 1000 μ l and precipitate DNA, centrifugal, abandon supernatant liquor, DNA precipitation is the washing with alcohol of 70% through volume fraction, after seasoning, solution (the OD that concentration is 20ng/ μ l is made with TE buffer solution
260/ OD
280=1.76 ~ 1.80) ,-20 DEG C save backup.
2, PCR random amplification sex determination interval screening sex specific molecular marker
The research of different lines bolti is shown, genetic linkage group LG1(Cnaani and Kocher, 2008; Lee et al., 2011; Lee et al., 2003), LG3(Campos-Ramos et al., 2001; Carrasco et al., 1999; Foresti et al., 1993; Harvey et al., 2003; Harvey et al., 2002; Ocalewicz et al., 2009) and LG23(Eshel et al., 2011; 2012) karyomit(e) corresponding to is all likely sex chromosome, but which LG correspondence karyomit(e) on earth, also do not come to a conclusion at present.Based on previous research and bolti genomic dna sequence, contriver to select in LG23 Random Design 32 pairs of primers on 3 adjacent Scaffold7,101 and 29 (be Scaffold716 respectively to, Scaffold1016 to, Scaffold2910 to), respectively pcr amplification is carried out to XX, XY, YY bolti genome pond, screening sex specific molecular marker.Found that, (upstream primer F1 sequence is as shown in SEQ ID No.1 for 1 pair of primer on Scaffold101, downstream primer R1 sequence is as shown in SEQ IDNo.2) there is obvious sex-specific, the X chromosome specific band of a long 1422bp can be amplified in XX and XY individuality, in XY and YY individuality, amplify the Y chromosome specific band of a long 982bp, and all the other 31 pairs of primers do not have sex-specific simultaneously.
3, the cloned and sequenced of sex specific molecular marker
Get the sepharose containing above-mentioned X chromosome specific band, Y chromosome specific band respectively, reclaim object fragment, be cloned into pMD-19T carrier, transformation of E. coli competent cell again, with blue hickie sieve method screening positive clone, picking hickie list bacterium colony, with PCR method qualification positive colony, then checks order.
The X chromosome specific molecular marker (sequence as shown in SEQ ID No.3, called after sdX) obtain order-checking and Y chromosome specific molecular marker (sequence as shown in SEQ ID No.4, called after sdY) carry out sequence alignment analysis.As shown in Figure 2, X mainly contains 3 places with the sequence difference of Y chromosome specific molecular marker to result: compared with X chromosome specific molecular marker, 1. Y chromosome specific molecular marker inserts 36bp sequence at 514bp place; 2. 4bp sequence has been lacked at 738bp place; 3. 472bp sequence has been lacked at 838bp place.
Two, the molecule marking method of bolti genetic sex qualification
The molecule marking method of bolti genetic sex qualification, comprises the following steps:
A.PCR clones: the fin ray of clip bolti to be measured, extracts genomic dna; With gained genomic dna for template, the upstream primer F1 shown in SEQ ID No.1 and the downstream primer R1 shown in SEQ ID No.2 is adopted to carry out pcr amplification; Pcr amplification parameter is: 94 DEG C of denaturations 3 minutes, and then 94 DEG C of sex change, 30 seconds, 58 DEG C annealing 30 seconds, 72 DEG C extend 30 seconds, totally 38 circulations, last 72 DEG C of extensions 10 minutes; Gained PCR primer carries out the agarose gel electrophoresis of 1.5%;
B. result judges: when only amplifying the X chromosome specific band (i.e. X chromosome specific molecular marker sdX of the present invention) of long 1422bp in the genomic dna from bolti to be measured, then judge that this bolti to be measured is the raun in heredity, sex chromosome genotype is XX; When only amplifying the Y chromosome specific band (i.e. Y chromosome specific molecular marker sdY of the present invention) of long 982bp in the genomic dna from bolti to be measured, then judge that this bolti to be measured is the supermale fish in heredity, sex chromosome genotype is YY; When the Y chromosome specific band of the X chromosome specific band and long 982bp that amplify long 1422bp in the genomic dna from bolti to be measured simultaneously, then judge that this bolti to be measured is the milter in heredity, sex chromosome genotype is XY.
Application aforesaid method carries out genetic sex qualification to 5 tail bolti raun XX, 5 tail bolti milter XY and 5 tail bolti supermale fish YY, the results are shown in Figure 2, the X chromosome specific band of long 1422bp has all only been amplified in all XX individualities, the Y chromosome specific band of long 982bp has all only been amplified in all YY individualities, and in XY individuality, amplified the X chromosome specific band of long 1422bp and the Y chromosome specific band of long 982bp simultaneously, consistent with notional result.
Application aforesaid method carries out genetic sex qualification to 19 tail bolti XX ♀ × XX ♂ filial generations, the results are shown in Figure 3, the X chromosome specific band of long 1422bp has all only been amplified in all boltis to be measured and contrast XX individuality, and amplified the X chromosome specific band of long 1422bp and the Y chromosome specific band of long 982bp in contrast XY individuality simultaneously, the Y chromosome specific band of long 1422bp has only been amplified in contrast YY individuality, illustrate that this 19 tail bolti to be measured is the XX raun in heredity, consistent with notional result.
Application aforesaid method carries out genetic sex qualification to 19 tail bolti XX ♀ × YY ♂ filial generations, the results are shown in Figure 4, the X chromosome specific band of long 1422bp and the Y chromosome specific band of long 1070bp have been amplified in all boltis to be measured and contrast XY individuality all simultaneously, and the X chromosome specific band of long 1422bp has only been amplified in contrast XX individuality, the Y chromosome specific band of long 982bp has only been amplified in contrast YY individuality, illustrate that this 19 tail bolti to be measured is the XY milter in heredity, consistent with notional result.
Application aforesaid method carries out genetic sex qualification to 19 tail bolti YY ♀ × YY ♂ filial generations, the results are shown in Figure 5, the Y chromosome specific band of long 1422bp has all only been amplified in all boltis to be measured and contrast YY individuality, and the X chromosome specific band of long 1422bp has only been amplified in contrast XX individuality, the X chromosome specific band of long 1422bp and the Y chromosome specific band of long 982bp have been amplified in contrast XY individuality simultaneously, illustrate that this 19 tail bolti to be measured is the YY supermale fish in heredity, consistent with notional result.
Three, the genetic linkage analysis in bolti sex specific molecular marker and sex determination site
Japanese strain bolti 3 the XX ♀ × XY ♂ introduced this laboratory normally cultivate family and carry out linkage analysis.The phenotypic sex of each individuality is confirmed by tissue slice, and genetic sex adopts bolti sex specific molecular marker sdX/sdY respectively, NtX1/NtY2, marker2 identify.Utilize JoinMAP4.0 to carry out the genetic linkage analysis of sex decision bit point (sex-determining loci, SD) and above sex specific molecular marker, the results are shown in Table 1 and Fig. 6.
The genetic linkage analysis in table 1 bolti sex specific molecular marker and sex determination site
Note: the individual individuality eliminating natural sex and reverse of restructuring; LOD threshold value is 3.0; Genetic distance and LOD value are calculated by Joinmap4.0.
As shown in Table 1, the genetic distance in bolti sex specific molecular marker sdX/sdY, NtX1/NtY2 and marker2 distance sex determination site is respectively 0,1.89,1.08cM, sex specific molecular marker sdX/sdY of the present invention be obviously better than another two groups of molecule markers.
Four, based on sex specific molecular marker, genetic sex qualification is carried out to the bolti of different sources
Adopt sex specific molecular marker sdX/sdY of the present invention, to from Hainan (HN1, HN2), the 98 tail boltis (62 tail phenotype rauns and 36 tail phenotype milters) of 4 different lines of Guangxi (THS) and Guangdong (NG) four tilapia nurseries carry out genetic sex qualification, and result is shown as 59 tail rauns and 39 tail milters.Carry out the matching analysis with phenotypic evaluation result, discovery has 5 coda gene type sexes and phenotypic sex qualification result is misfitted, and wherein 4 tail phenotype rauns are accredited as XY milter, and 1 tail phenotype milter is accredited as XX raun; The bolti of 4 different lines, the identical rate of genetic sex and phenotypic sex is 90% ~ 100%, and what rate of coincideing was minimum is HN2 family, and the highest is HN1 family (table 2), and the reason misfitted may be caused by tilapia generation sex reversal.From the above results, sex specific molecular marker sdX/sdY of the present invention is applicable to the bolti of this Laboratory culture and the bolti of 4 different sources strains, has universality.
The genetic sex qualification result of table 24 different sources family boltis
| Phenotypic sex | HN1(28) | HN2(22) | THS(25) | NG(23) |
| Raun | (19♀:0♂) | (13♀:1♂) | (13♀:2♂) | (13♀:1♂) |
| Milter | (0♀:9♂) | (1♀:7♂) | (0♀:10♂) | (0♀:9♂) |
| Raun accuracy rate | 100% | 93% | 87% | 93% |
| Milter accuracy rate | 100% | 88% | 100% | 100% |
| Average Accuracy | 100% | 90% | 93% | 96% |
Five, the method for supermale bolti is produced based on sex specific molecular marker
Carry out the qualification of bolti genetic sex based on sex specific molecular marker sdX/sdY of the present invention, the method for producing supermale bolti comprises the following steps:
A. bolti raun XX and milter XY post-coitum generation are carried out estradiol dipping bath in hatching latter 5 days, the molecule marking method adopting bolti genetic sex of the present invention to identify, filters out the conversion raun that sex chromosome genotype is XY from sexual maturity female individuals;
B. XY is transformed raun and the mating of bolti XY milter, the molecule marking method adopting bolti genetic sex of the present invention to identify, from sexual maturity offspring, filter out the supermale fish that sex chromosome genotype is YY; Another XY is transformed raun with bolti XY milter post-coitum for carrying out estradiol dipping bath in hatching latter 5 days, adopt the molecule marking method of bolti genetic sex qualification, from sexual maturity female individuals, filter out the conversion raun that sex chromosome genotype is YY;
C. YY supermale fish and YY are transformed raun mating, obtain full YY supermale fish;
D. YY supermale fish offspring is carried out feminized treatment, obtain YY and transform raun;
E. repeating step C and D, transforms raun mating by YY supermale fish and YY and produces YY supermale fish.
After employing aforesaid method ensures to continue to obtain YY supermale fish, by YY supermale fish and the mating of bolti XX raun, complete male marketable fish can be produced in a large number.
What finally illustrate is, above preferred embodiment is only in order to illustrate technical scheme of the present invention and unrestricted, although by above preferred embodiment to invention has been detailed description, but those skilled in the art are to be understood that, various change can be made to it in the form and details, and not depart from claims of the present invention limited range.
Claims (3)
1. Y-chromosome specific molecular marker of Nile tilapia, is characterized in that, nucleotide sequence is as shown in SEQ ID No.4.
2. utilize Y-chromosome specific molecular marker of Nile tilapia described in claim 1 to carry out the method for bolti genetic sex qualification, comprise the following steps:
A.PCR increases: the fin ray getting bolti to be measured, extracts genomic dna; With gained genomic dna for template, adopt the upstream primer shown in SEQ ID No.1 and the downstream primer shown in SEQ ID No.2, pcr amplification bolti sex specific molecular marker: the nucleotide sequence of X chromosome specific molecular marker is as shown in SEQ ID No.3, and the nucleotide sequence of Y chromosome specific molecular marker is as shown in SEQ ID No.4;
B. result judges: from the genomic dna of bolti to be measured, only amplify X chromosome specific molecular marker, judge that this bolti to be measured is the raun in heredity, sex chromosome genotype is XX; From the genomic dna of bolti to be measured, only amplify Y chromosome specific molecular marker, judge that this bolti to be measured is the supermale fish in heredity, sex chromosome genotype is YY; From the genomic dna of bolti to be measured, amplify X chromosome specific molecular marker and Y chromosome specific molecular marker, judge that this bolti to be measured is the milter in heredity, sex chromosome genotype is XY simultaneously.
3. adopt the method for bolti genetic sex qualification described in claim 2 to produce the method for supermale fish, comprise the following steps:
A. in the post-coitum of bolti raun XX and milter XY generation, is carried out feminized treatment, adopt the method for bolti genetic sex qualification described in claim 2, filter out the conversion raun that sex chromosome genotype is XY;
B. XY is transformed raun and the mating of bolti XY milter, adopt the method for bolti genetic sex qualification described in claim 2, from offspring, filter out the supermale fish that sex chromosome genotype is YY; Separately get XY to transform raun and bolti XY milter post-coitum generation and carry out feminized treatment, adopt the method that described in claim 2, bolti genetic sex is identified, filter out the conversion raun that sex chromosome genotype is YY;
C. YY supermale fish and YY are transformed raun mating, obtain full YY supermale fish;
D. YY supermale fish offspring is carried out feminized treatment, obtain YY and transform raun;
E. repeating step C and D, transforms raun mating by YY supermale fish and YY and produces YY supermale fish.
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| CN110250116A (en) * | 2019-08-05 | 2019-09-20 | 广东海洋大学 | A kind of method of quickly breeding YY supermale bolti |
| CN112655600A (en) * | 2019-12-06 | 2021-04-16 | 海南勤富实业有限公司 | Method for quickly cultivating tilapia mossambica in early stage |
| CN110938626B (en) * | 2019-12-12 | 2023-07-18 | 西南大学 | Specific molecular marker of sex chromosome of Oreotilapia mossambica, genetic sex identification based on molecular marker and production method of unisex fish |
| CN116042863A (en) * | 2023-03-13 | 2023-05-02 | 内江师范学院 | Method for detecting nile tilapia allele expression level based on double-fluorescence-labeled probe |
| CN116804225B (en) * | 2023-07-24 | 2024-02-13 | 西部(重庆)科学城种质创制大科学中心 | Ji Shi tilapia sex chromosome specific molecular marker, detection primer, kit and application thereof |
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| CN101932641A (en) * | 2008-01-30 | 2010-12-29 | 昭和电工株式会社 | Transparent composite |
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| A Genetic Linkage Map of a Cichlid Fish,the Tilapia(Oreochromis niloticus);Thomas D.Kocher et al.;《Genetics》;19980331;第148卷;第1225-1232页 * |
| Isolation,sequence analysis,and characterization of androgen receptor in Southern catfish,Silurus meridionalis;B.F.Huang et al.;《Fish Physiol Biochem》;20101229;第37卷;第593-601页 * |
| Linkage and Physical Mapping of Sex Region on LG23 of Nile Tilapia(Oreochromis niloticus);O.Eshel et al.;《G3》;20120131;第2卷;摘要，第39页右栏第2段 * |
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