In this study, we conducted a cross of white crucian carp （♀）xred crucian carp （♂） （WR）, and characterized the morphology, reproduction and genetics of the progeny. Different from parents, WR with the gray col...In this study, we conducted a cross of white crucian carp （♀）xred crucian carp （♂） （WR）, and characterized the morphology, reproduction and genetics of the progeny. Different from parents, WR with the gray color showed the hybrid morphological traits of both parents. WR possessed normal gonads producing mature eggs or sperm, and exhibited high fertilization rate （90.2%） and high hatchery rate （80.5%）, which contributed to produce and enlarge the population. WR with the same DNA content as parents was a diploid fish with 100 chromosomes （2n=100）. Amplified ITS of 45S rDNA, in WR the sequences consisting of 884 bp bases of the entire ITS-1 region, 5.8 S region, and entire ITS-2 region. The sequences showed high similarity between WR and its parents and leaned towards male inheritance. In WR, NTS of 5S rDNA consisted of three length types with total 654 bp bases. From sequence analysis of NTS, WR shared 94.2% and 95.1% similarities with their female and male parent, respectively. Sequence analysis of ITS and NTS revealed that there existed recombination and variation in the hybrid progeny, which was the genetic base for adaptation and speciation. In conclusion, we obtained WR from hybridization and it exhibited hybrid traits in morphology and variation in genetic composition showing essential difference with its parents. The obtainment of WR has important significance in fish genetic breeding.展开更多
The white crucian carp(Carassius auratus cuvieri,WCC) not only is one of the most economically important fish in Asia,characterized by strong reproductive ability and rapid growth rates,but also represents a good germ...The white crucian carp(Carassius auratus cuvieri,WCC) not only is one of the most economically important fish in Asia,characterized by strong reproductive ability and rapid growth rates,but also represents a good germplasm to produce hybrid progenies with heterosis.Gene knockout technique provides a safe and acceptant way for fish breeding.Achieving gene knockout in WCC and its hybrid progeny will be of great importance for both genetic studies and hybridization breeding.Tyrosinase(TYR) is a key enzyme in melanin synthesis.Depletion of tyr in zebrafish and mice results in mosaic pigmentation or total albinism.Here,we successfully used CRISPR-Cas9 to target tyr in WCC and its hybrid progeny(WR) derived from the cross of WCC(♀) and red crucian carp(Carassius auratus red var.,RCC,♂).The level of TYR protein was significantly reduced in mutant WCC.Both the mutant WCC and the mutant WR showed different degrees of melanin reduction compared with the wild-type sibling control fish,resulting from different mutation efficiency ranging from 60% to 90%.In addition,the transcriptional expression profiles of a series of pivotal pigment synthesis genes,i.e.tyrp1,mitfa,mitfb,dct and sox10,were down-regulated in tyr-CRISPR WCC,which ultimately caused a reduction in melanin synthesis.These results demonstrated that tyr plays a key role in melanin synthesis in WCC and WR,and CRISPR-Cas9 is an effective tool for modifying the genome of economical fish.Furthermore,the tyr-CRISPR models could be valuable in understanding fundamental mechanisms of pigment formation in non-model fish.展开更多
Triploid carp(100%)with 150(3n=150)chromosomes were obtained by crossing the females of improved tetraploid hybrids(♀, 4n=200)of red crucian carp(♀)×common carp(♂)with the males of diploid yellow river carp(♂...Triploid carp(100%)with 150(3n=150)chromosomes were obtained by crossing the females of improved tetraploid hybrids(♀, 4n=200)of red crucian carp(♀)×common carp(♂)with the males of diploid yellow river carp(♂,2n=100).The crosses yielded transgenic triploid carp(positive triploid fish,44.2%of the progeny)and non-transgenic triploid carp(negative triploid fish). Histological examination of the gonads of 24-month-old positive triploid fish suggested they were sterile and the fish were not able to produce mature gametes during the breeding season.Morphologically,both the positive and negative triploid fish were similar.They had a spindle-shaped,laterally compressed,steel grey body with two pairs of barbells.Most of the quantifiable traits of the triploid carp were intermediate between those of the two parents.The positive and negative triploid fish were raised in the same pond for 2 years.The mean body weight of the positive triploid fish was 2.3 times higher than the negative triploid fish.The weight of the largest positive triploid fish was 2.91 times higher than that of the largest negative triploid fish.Thus,we produced fast-growing transgenic triploid carp that have a reduced ecological risk because of their inability to mate and produce progeny.展开更多
In terms of taxonomic status,common carp(Cyprinus carpio,Cyprininae)and crucian carp(Carassius auratus,Cyprininae)are different species;however,in this study,a newborn homodiploid crucian carp-like fish(2n=100)(2nNCRC...In terms of taxonomic status,common carp(Cyprinus carpio,Cyprininae)and crucian carp(Carassius auratus,Cyprininae)are different species;however,in this study,a newborn homodiploid crucian carp-like fish(2n=100)(2nNCRC)lineage(F1–F3)was established from the interspecific hybridization of female common carp(2n=100)×male blunt snout bream(Megalobrama amblycephala,Cultrinae,2n=48).The phenotypes and genotypes of 2 n NCRC differed from those of its parents but were closely related to those of the existing diploid crucian carp.We further sequenced the whole mitochondrial(mt)genomes of the 2n NCRC lineage from F1 to F3.The paternal mt DNA fragments were stably embedded in the mt-genomes of F1–F3 generations of 2n NCRC to form chimeric DNA fragments.Along with this chimeric process,numerous base sites of F1–F3 generations of 2 n NCRC underwent mutations.Most of these mutation sites were consistent with the existing diploid crucian carp.Moreover,the mt DNA organization and nucleotide composition of 2n NCRC were more similar to those of the existing diploid crucian carp than those of the parents.The inheritable chimeric DNA fragments and mutant loci in the mt-genomes of different generations of 2nNCRC provided important evidence of the mt DNA change process in the newborn lineage derived from hybridization of different species.Our findings demonstrated for the first time that the paternal mt DNA were transmitted into the mt-genomes of homodiploid lineage,which provided new insights into the existence of paternal mt DNA in the mt DNA inheritance.展开更多
Common carp Cyprinus carpio genomic DNA repetitive sequence CR1 has been DIG-labeled and hybridized in situ against chromosomes of red common carp ( Cyprinus carpio L. Xingguo red var.). It is found that the repetitiv...Common carp Cyprinus carpio genomic DNA repetitive sequence CR1 has been DIG-labeled and hybridized in situ against chromosomes of red common carp ( Cyprinus carpio L. Xingguo red var.). It is found that the repetitive sequence CR1 is mainly localized at the cen-tromeric regions of chromosomes of the red common carp. The application of the展开更多
Mitochondrial DNA fragment of cytb was compared in all species Carassius auratus complex and three forms of ornamental goldfish. It is shown that the phylogenetic relationships between complex species correspond to th...Mitochondrial DNA fragment of cytb was compared in all species Carassius auratus complex and three forms of ornamental goldfish. It is shown that the phylogenetic relationships between complex species correspond to the existing views, based on mtDNA data and geographical distribution. All forms of ornamental goldfish have a monophyletic origin from Chinese goldfish C. auratus auratus. The analysis showed that three nuclear genes (rps7, GH1 and Rh) in the two forms of ornamental goldfish (Oriental twintail goldfish and Chinese Ranchu) were almost identical C. auratus auratus genes, while all three gene in another more simple form of goldfish (common goldfish) were highly homologous to carp Cyprinus carpio nuclear genes. The obtained data suggested that in the history of aquarium goldfish breeding occurred the stage of distant hybridization between goldfish and common carp. Subsequently, the nuclear genomes of some ornamental forms could be enriched by goldfish genes (a relatively recent form as Oriental twintail goldfish and Chinese Ranchu) or common carp genes (the simplest and most ancient forms like common goldfish) as a result of multidirectional breeding and selection of aquarium goldfish various forms.展开更多
采用 DNA 杂交的方法,对异育银鲫及其人工杂合种进行了外源 DNA 的检测分析,发现两个雌核发育系的异育银鲫及其人工杂合种的 DNA 与红鲤 DNA 片段间的分子杂交均有阳性斑点出现,这表明,在异育银鲫及其人工杂合种产生过程中,父本(红鲤)的...采用 DNA 杂交的方法,对异育银鲫及其人工杂合种进行了外源 DNA 的检测分析,发现两个雌核发育系的异育银鲫及其人工杂合种的 DNA 与红鲤 DNA 片段间的分子杂交均有阳性斑点出现,这表明,在异育银鲫及其人工杂合种产生过程中,父本(红鲤)的 DNA 片段的确可以随机地掺入到母本细胞的 DNA 中,从而产生了异精效应,使其子代生长加快,并出现父本性状。展开更多
基金supported by Major International Cooperation Projects of the National Natural Science Foundation of China(31210103918)the National Natural Science Foundation of China(31201980,31430088 and 31272651)+5 种基金the National High Technology Research and Development Program of China(2011AA100403)the Natural Science Foundation of Hunan Province,China(14JJ3072)Science-Technology Foundation of Hunan Province,China(2014FJ3084)Research Foundation of Education Bureau of Hunan Province,China(13C523)the Cooperative Innovation Center of Engineering and New Products for Developmental Biology of Hunan Province(20134486)the Construct Program of the Key Discipline in Hunan Province,and Construct Program of the National Key Discipline
文摘In this study, we conducted a cross of white crucian carp （♀）xred crucian carp （♂） （WR）, and characterized the morphology, reproduction and genetics of the progeny. Different from parents, WR with the gray color showed the hybrid morphological traits of both parents. WR possessed normal gonads producing mature eggs or sperm, and exhibited high fertilization rate （90.2%） and high hatchery rate （80.5%）, which contributed to produce and enlarge the population. WR with the same DNA content as parents was a diploid fish with 100 chromosomes （2n=100）. Amplified ITS of 45S rDNA, in WR the sequences consisting of 884 bp bases of the entire ITS-1 region, 5.8 S region, and entire ITS-2 region. The sequences showed high similarity between WR and its parents and leaned towards male inheritance. In WR, NTS of 5S rDNA consisted of three length types with total 654 bp bases. From sequence analysis of NTS, WR shared 94.2% and 95.1% similarities with their female and male parent, respectively. Sequence analysis of ITS and NTS revealed that there existed recombination and variation in the hybrid progeny, which was the genetic base for adaptation and speciation. In conclusion, we obtained WR from hybridization and it exhibited hybrid traits in morphology and variation in genetic composition showing essential difference with its parents. The obtainment of WR has important significance in fish genetic breeding.
基金supported by the National Natural Science Foundation of China (31430088, 31730098)the earmarked fund for China Agriculture Research System (CARS-45)+2 种基金the Cooperative Innovation Center of Engineering and New Products for Developmental Biology of Hunan Province (20134486)the Natural Science Foundation of Hunan Province (14JJ2062)the Research Foundation of Education Bureau of Hunan Province, China (16B160)
文摘The white crucian carp(Carassius auratus cuvieri,WCC) not only is one of the most economically important fish in Asia,characterized by strong reproductive ability and rapid growth rates,but also represents a good germplasm to produce hybrid progenies with heterosis.Gene knockout technique provides a safe and acceptant way for fish breeding.Achieving gene knockout in WCC and its hybrid progeny will be of great importance for both genetic studies and hybridization breeding.Tyrosinase(TYR) is a key enzyme in melanin synthesis.Depletion of tyr in zebrafish and mice results in mosaic pigmentation or total albinism.Here,we successfully used CRISPR-Cas9 to target tyr in WCC and its hybrid progeny(WR) derived from the cross of WCC(♀) and red crucian carp(Carassius auratus red var.,RCC,♂).The level of TYR protein was significantly reduced in mutant WCC.Both the mutant WCC and the mutant WR showed different degrees of melanin reduction compared with the wild-type sibling control fish,resulting from different mutation efficiency ranging from 60% to 90%.In addition,the transcriptional expression profiles of a series of pivotal pigment synthesis genes,i.e.tyrp1,mitfa,mitfb,dct and sox10,were down-regulated in tyr-CRISPR WCC,which ultimately caused a reduction in melanin synthesis.These results demonstrated that tyr plays a key role in melanin synthesis in WCC and WR,and CRISPR-Cas9 is an effective tool for modifying the genome of economical fish.Furthermore,the tyr-CRISPR models could be valuable in understanding fundamental mechanisms of pigment formation in non-model fish.
基金supported by the National Key Basic Research Program of China (2007CB109206)the National Special Fund for Research in Public Welfare Sector (200903046-08)
文摘Triploid carp(100%)with 150(3n=150)chromosomes were obtained by crossing the females of improved tetraploid hybrids(♀, 4n=200)of red crucian carp(♀)×common carp(♂)with the males of diploid yellow river carp(♂,2n=100).The crosses yielded transgenic triploid carp(positive triploid fish,44.2%of the progeny)and non-transgenic triploid carp(negative triploid fish). Histological examination of the gonads of 24-month-old positive triploid fish suggested they were sterile and the fish were not able to produce mature gametes during the breeding season.Morphologically,both the positive and negative triploid fish were similar.They had a spindle-shaped,laterally compressed,steel grey body with two pairs of barbells.Most of the quantifiable traits of the triploid carp were intermediate between those of the two parents.The positive and negative triploid fish were raised in the same pond for 2 years.The mean body weight of the positive triploid fish was 2.3 times higher than the negative triploid fish.The weight of the largest positive triploid fish was 2.91 times higher than that of the largest negative triploid fish.Thus,we produced fast-growing transgenic triploid carp that have a reduced ecological risk because of their inability to mate and produce progeny.
基金supported by the National Natural Science Foundation of China(31802287,31730098,31430088)the China Postdoctoral Science Foundation(2018M632969)+3 种基金the earmarked fund for China Agriculture Research System(CARS-45)the Key Research and Development Program of Hunan Province(2018NK2072)Hunan Provincial Natural Science and Technology Major Project(2017NK1031)the Cooperative Innovation Center of Engineering and New Products for Developmental Biology of Hunan Province(20134486)
文摘In terms of taxonomic status,common carp(Cyprinus carpio,Cyprininae)and crucian carp(Carassius auratus,Cyprininae)are different species;however,in this study,a newborn homodiploid crucian carp-like fish(2n=100)(2nNCRC)lineage(F1–F3)was established from the interspecific hybridization of female common carp(2n=100)×male blunt snout bream(Megalobrama amblycephala,Cultrinae,2n=48).The phenotypes and genotypes of 2 n NCRC differed from those of its parents but were closely related to those of the existing diploid crucian carp.We further sequenced the whole mitochondrial(mt)genomes of the 2n NCRC lineage from F1 to F3.The paternal mt DNA fragments were stably embedded in the mt-genomes of F1–F3 generations of 2n NCRC to form chimeric DNA fragments.Along with this chimeric process,numerous base sites of F1–F3 generations of 2 n NCRC underwent mutations.Most of these mutation sites were consistent with the existing diploid crucian carp.Moreover,the mt DNA organization and nucleotide composition of 2n NCRC were more similar to those of the existing diploid crucian carp than those of the parents.The inheritable chimeric DNA fragments and mutant loci in the mt-genomes of different generations of 2nNCRC provided important evidence of the mt DNA change process in the newborn lineage derived from hybridization of different species.Our findings demonstrated for the first time that the paternal mt DNA were transmitted into the mt-genomes of homodiploid lineage,which provided new insights into the existence of paternal mt DNA in the mt DNA inheritance.
文摘Common carp Cyprinus carpio genomic DNA repetitive sequence CR1 has been DIG-labeled and hybridized in situ against chromosomes of red common carp ( Cyprinus carpio L. Xingguo red var.). It is found that the repetitive sequence CR1 is mainly localized at the cen-tromeric regions of chromosomes of the red common carp. The application of the
文摘Mitochondrial DNA fragment of cytb was compared in all species Carassius auratus complex and three forms of ornamental goldfish. It is shown that the phylogenetic relationships between complex species correspond to the existing views, based on mtDNA data and geographical distribution. All forms of ornamental goldfish have a monophyletic origin from Chinese goldfish C. auratus auratus. The analysis showed that three nuclear genes (rps7, GH1 and Rh) in the two forms of ornamental goldfish (Oriental twintail goldfish and Chinese Ranchu) were almost identical C. auratus auratus genes, while all three gene in another more simple form of goldfish (common goldfish) were highly homologous to carp Cyprinus carpio nuclear genes. The obtained data suggested that in the history of aquarium goldfish breeding occurred the stage of distant hybridization between goldfish and common carp. Subsequently, the nuclear genomes of some ornamental forms could be enriched by goldfish genes (a relatively recent form as Oriental twintail goldfish and Chinese Ranchu) or common carp genes (the simplest and most ancient forms like common goldfish) as a result of multidirectional breeding and selection of aquarium goldfish various forms.
文摘采用 DNA 杂交的方法,对异育银鲫及其人工杂合种进行了外源 DNA 的检测分析,发现两个雌核发育系的异育银鲫及其人工杂合种的 DNA 与红鲤 DNA 片段间的分子杂交均有阳性斑点出现,这表明,在异育银鲫及其人工杂合种产生过程中,父本(红鲤)的 DNA 片段的确可以随机地掺入到母本细胞的 DNA 中,从而产生了异精效应,使其子代生长加快,并出现父本性状。
