长鳍鲤、锦鲤和龙凤鲤的遗传多样性研究
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摘要
本研究包括三部分,第一部分以龙凤鲤(♂)×锦鲤(♀)、龙凤鲤(♂)×龙凤鲤(♀)为亲本,得到两组子代龙凤鲤(93尾),测量这两组子代龙凤鲤的10个主要形态性状和20个框架结构参数,研究亲本对子代形态的影响;第二部分以长鳍鲤、锦鲤和龙凤鲤共76尾个体为研究素材,从鲤鱼微卫星引物中筛选20对引物,利用微卫星分子标记方法,分析基因组DNA在20个微卫星位点上的遗传差异;第三部分以长鳍鲤8尾、锦鲤8尾和龙凤鲤个体10尾为研究对象,利用线粒体DNA D-loop区分子标记方法,研究这3种鲤鱼在线粒体DNA非编码区的遗传差异和亲缘关系,主要研究结果如下:
1、两组子代30个形态指标通过均值t检验、主成分分析、判别分析和差异系数检验方法分析得到,两组子代只在体厚/体宽、胸鳍基起点至头部背侧末端BC/体长、背鳍基起点至臀鳍基起点EF/体长和背鳍基起点至臀鳍基末端EH/体长四个指标具有显著差异(P<0.05),其余指标均没有显著差异;主成分分析得到9个主成分,累积贡献率为81.11%,主成分分析图显示彼此之间没有明显的偏离态势;判别分析得到以体厚/体长、吻长/体长、眼间距/体长、胸鳍基起点至腹鳍基起点BD/体长、胸鳍基起点至头部背侧末端BC/体长和腹鳍基起点至头部背侧末端DC/体长6个指标为分析因子的判别方程,两组子代的判别准确率为78.5%;差异系数检验发现30个形态指标的差异系数值均小于1.28,没有达到划分亚种的标准。从总体判断,由龙凤鲤(♂)×龙凤鲤(♀)的子代和由龙凤鲤(♂)×锦鲤(♀)的子代在形态上具有高度的相似性。
2、使用20对微卫星引物对长鳍鲤、锦鲤和龙凤鲤的遗传多样性进行研究,发现3个群体在20个位点上分别检测到108、164、154个等位基因,有效等位基因数分别为4.496、5.695和5.506,遗传多样性分别为0.764、0.803和0.799,期望杂合度分别为0.769、0.806和0.801,多态信息含量(PIC)分别为0.703、0.761和0.757,总体遗传多样性水平较高,其中锦鲤的遗传多样性最高,龙凤鲤次之,长鳍鲤最低。Hardy-Weinberg平衡检验发现,3个群体共有26个基因座表现为极显著的遗传不平衡(P<0.01),偏离指数分析显示:3个群体在20个微卫星位点上大部分处于杂合子过剩状态。平均遗传分化系数为7.1%,基因流Nm的平均值为3.5895,3个群体间基因流动性较大。
3、对长鳍鲤、锦鲤和龙凤鲤的线粒体DNA D-loop区序列进行遗传结构和亲缘关系分析,得到长度为386 bp的分析片段,该片段富含A+T,G+C含量较少,共发现17个变异位点,约占分析总位点的4.66%,其中单一多态位点7个,简约信息位点10个,观测到一个碱基缺失及174 bp处的A碱基插入。共发现10种单倍型,其中2种单倍型为群体共享,其他均为各群体所特有,群体平均单倍型变异为0.708±0.095,3个群体的平均核苷酸变异和核苷酸多态性分别为3.966和0.0103,遗传聚类分析显示,龙凤鲤和锦鲤遗传距离近,而与长鳍鲤的遗传距离较远,说明在3个群体中龙凤鲤在很多遗传性状上与锦鲤相似,锦鲤的遗传多样性指标高于长鳍鲤和龙凤鲤,结果与微卫星分析显示的相同。
There were three parts of this article,the first part was to find the relationships between two groups of Long-feng carp(93),which crossed by two different parents,one was Long-feng(♂)×Long-feng carp(♀) ,the other one was Long-feng(♂)×Koi carp(♀). Ten traditional morphological data and twenty truss network data were combined to conduct a multivariate analysis to study the influence of different parents.the second part used twenty microsatellite primers to find the genetic diversity of three groups of carps----- the Long-fin carp,Koi carp and Long-feng carp(total sample sizes was 76 ). In the third part ,choosed 8 of Long-fin carp,8 of Koi carp and 10 of Long-feng carp as samples to find the difference of these 3 carps in genomic DNA and the genetic relationships by mitochondrial DNA D-loop methord .and the results were as follows:
1. Except the traits of body width, distance between the origin of pectoral fin and terminal of head back(BC),the origin of dorsal fin and origin of anal fin(EF) and the origin of dorsal fin and terminal of anal fin(EH),There had no significant difference(P<0.05) between the two groups of the morphological traits tested by the mean value. principal component analysis showed that all traits could be divided into 9 kinds of factors ,and the cumulative contribution ratio was 81.11%,there had no obviously seprate trend in the two populations,the discriminant analysis indicated that, by six traits—body width,snout length,eyes distances,distance between the origin of pectoral fin and origin of pelvic fin(BD),distance between the origin of pectoral fin and terminal of head back(BC) and distance between the origin of pelvic fin and terminal of head back( DC) could get two discriminate functions, the total discriminant accuracy was 78.5% .the difference coefficient test found that all of the 30 difference values were less than 1.28,did not met the criteria into sub-species,the conclusion was that there had high similarity between the two study groups although they had different parents.
2. Twenty microsatellite primers were use to study the genetic diversity of Long-fin carp,Koi carp and Long-feng carp,The results were as follows: 108,164 and 154 alleles were respectively detected on the 20 loci in the three carp populations.the effective number of alleles in each population was 4.496,5.965 and 5.506 , the genetic diversity were 0.764,0.803 and 0.799, the expected heterozygosity were 0.769,0.806 and 0.801, the polymorphism information content (PIC)were 0.703,0.761 and 0.757 respectively, the three carp groups had a higher genetic diversity levels overall.in which Koi carp was the highest,Long-feng second,then the Long-fin carp.Hardy-Weinberg balance test revealed that there had 26 loci in 3 carp groups showed significant genetic imbalance(P<0.01),the deviation index analysis concluded that,most of the 20 loci were heterozygote excess,The average genetic differentiation coefficient was 7.1%,average gene flow was 3.5895,had a relatively high gene flow in 3 carp groups.
3. In the present study ,population structure and the genetic relationship of three carp groups,Long-fin carp, Koi carp and Long-feng carp ,were investigated using mitochondrial DNA sequence data analysis.A total of 386bp fragment were analysed,in which was rich in A+T,less in G+C, 17 vavialbe sites were found, in which had 7singleton variable sites , 10 parsimony informative sites , 1 site with missing data ,and one site with A base insert,10 haplotypes were revealed,2 of them were shared by the group,others were specific to each group,the average haplotype diversity of the groups was 0.708±0.095,the average number of nucleotide differences was 3.966 and the nucleotide diversity was 0.013,Genetic cluster analysis showed that ,the Long-feng carp had a closer genetic distance with Koi carp,the conclusion was that in the three carp populations ,Long-feng carp was much similar to Koi carp,the Koi carp `s genetic diversity index were higher than Long-feng carp and Long-fin carp,which was as the same as the results of microsatellite analysis.
1、两组子代30个形态指标通过均值t检验、主成分分析、判别分析和差异系数检验方法分析得到,两组子代只在体厚/体宽、胸鳍基起点至头部背侧末端BC/体长、背鳍基起点至臀鳍基起点EF/体长和背鳍基起点至臀鳍基末端EH/体长四个指标具有显著差异(P<0.05),其余指标均没有显著差异;主成分分析得到9个主成分,累积贡献率为81.11%,主成分分析图显示彼此之间没有明显的偏离态势;判别分析得到以体厚/体长、吻长/体长、眼间距/体长、胸鳍基起点至腹鳍基起点BD/体长、胸鳍基起点至头部背侧末端BC/体长和腹鳍基起点至头部背侧末端DC/体长6个指标为分析因子的判别方程,两组子代的判别准确率为78.5%;差异系数检验发现30个形态指标的差异系数值均小于1.28,没有达到划分亚种的标准。从总体判断,由龙凤鲤(♂)×龙凤鲤(♀)的子代和由龙凤鲤(♂)×锦鲤(♀)的子代在形态上具有高度的相似性。
2、使用20对微卫星引物对长鳍鲤、锦鲤和龙凤鲤的遗传多样性进行研究,发现3个群体在20个位点上分别检测到108、164、154个等位基因,有效等位基因数分别为4.496、5.695和5.506,遗传多样性分别为0.764、0.803和0.799,期望杂合度分别为0.769、0.806和0.801,多态信息含量(PIC)分别为0.703、0.761和0.757,总体遗传多样性水平较高,其中锦鲤的遗传多样性最高,龙凤鲤次之,长鳍鲤最低。Hardy-Weinberg平衡检验发现,3个群体共有26个基因座表现为极显著的遗传不平衡(P<0.01),偏离指数分析显示:3个群体在20个微卫星位点上大部分处于杂合子过剩状态。平均遗传分化系数为7.1%,基因流Nm的平均值为3.5895,3个群体间基因流动性较大。
3、对长鳍鲤、锦鲤和龙凤鲤的线粒体DNA D-loop区序列进行遗传结构和亲缘关系分析,得到长度为386 bp的分析片段,该片段富含A+T,G+C含量较少,共发现17个变异位点,约占分析总位点的4.66%,其中单一多态位点7个,简约信息位点10个,观测到一个碱基缺失及174 bp处的A碱基插入。共发现10种单倍型,其中2种单倍型为群体共享,其他均为各群体所特有,群体平均单倍型变异为0.708±0.095,3个群体的平均核苷酸变异和核苷酸多态性分别为3.966和0.0103,遗传聚类分析显示,龙凤鲤和锦鲤遗传距离近,而与长鳍鲤的遗传距离较远,说明在3个群体中龙凤鲤在很多遗传性状上与锦鲤相似,锦鲤的遗传多样性指标高于长鳍鲤和龙凤鲤,结果与微卫星分析显示的相同。
There were three parts of this article,the first part was to find the relationships between two groups of Long-feng carp(93),which crossed by two different parents,one was Long-feng(♂)×Long-feng carp(♀) ,the other one was Long-feng(♂)×Koi carp(♀). Ten traditional morphological data and twenty truss network data were combined to conduct a multivariate analysis to study the influence of different parents.the second part used twenty microsatellite primers to find the genetic diversity of three groups of carps----- the Long-fin carp,Koi carp and Long-feng carp(total sample sizes was 76 ). In the third part ,choosed 8 of Long-fin carp,8 of Koi carp and 10 of Long-feng carp as samples to find the difference of these 3 carps in genomic DNA and the genetic relationships by mitochondrial DNA D-loop methord .and the results were as follows:
1. Except the traits of body width, distance between the origin of pectoral fin and terminal of head back(BC),the origin of dorsal fin and origin of anal fin(EF) and the origin of dorsal fin and terminal of anal fin(EH),There had no significant difference(P<0.05) between the two groups of the morphological traits tested by the mean value. principal component analysis showed that all traits could be divided into 9 kinds of factors ,and the cumulative contribution ratio was 81.11%,there had no obviously seprate trend in the two populations,the discriminant analysis indicated that, by six traits—body width,snout length,eyes distances,distance between the origin of pectoral fin and origin of pelvic fin(BD),distance between the origin of pectoral fin and terminal of head back(BC) and distance between the origin of pelvic fin and terminal of head back( DC) could get two discriminate functions, the total discriminant accuracy was 78.5% .the difference coefficient test found that all of the 30 difference values were less than 1.28,did not met the criteria into sub-species,the conclusion was that there had high similarity between the two study groups although they had different parents.
2. Twenty microsatellite primers were use to study the genetic diversity of Long-fin carp,Koi carp and Long-feng carp,The results were as follows: 108,164 and 154 alleles were respectively detected on the 20 loci in the three carp populations.the effective number of alleles in each population was 4.496,5.965 and 5.506 , the genetic diversity were 0.764,0.803 and 0.799, the expected heterozygosity were 0.769,0.806 and 0.801, the polymorphism information content (PIC)were 0.703,0.761 and 0.757 respectively, the three carp groups had a higher genetic diversity levels overall.in which Koi carp was the highest,Long-feng second,then the Long-fin carp.Hardy-Weinberg balance test revealed that there had 26 loci in 3 carp groups showed significant genetic imbalance(P<0.01),the deviation index analysis concluded that,most of the 20 loci were heterozygote excess,The average genetic differentiation coefficient was 7.1%,average gene flow was 3.5895,had a relatively high gene flow in 3 carp groups.
3. In the present study ,population structure and the genetic relationship of three carp groups,Long-fin carp, Koi carp and Long-feng carp ,were investigated using mitochondrial DNA sequence data analysis.A total of 386bp fragment were analysed,in which was rich in A+T,less in G+C, 17 vavialbe sites were found, in which had 7singleton variable sites , 10 parsimony informative sites , 1 site with missing data ,and one site with A base insert,10 haplotypes were revealed,2 of them were shared by the group,others were specific to each group,the average haplotype diversity of the groups was 0.708±0.095,the average number of nucleotide differences was 3.966 and the nucleotide diversity was 0.013,Genetic cluster analysis showed that ,the Long-feng carp had a closer genetic distance with Koi carp,the conclusion was that in the three carp populations ,Long-feng carp was much similar to Koi carp,the Koi carp `s genetic diversity index were higher than Long-feng carp and Long-fin carp,which was as the same as the results of microsatellite analysis.
引文
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