【作者】 林凯东；

【导师】 罗琛；

【作者基本信息】 湖南师范大学 ， 动物学， 2003， 硕士

【摘要】 本研究运用微卫星DNA-聚合酶链反应-聚丙烯酰胺凝胶电泳基因分型技术，从28对已发表的普通鲤鱼的微卫星引物中，筛选出能用于草鱼基因组分析的微卫星引物，然后用这些引物对湘江野生草鱼群体和通过抑制第一次卵裂获得的雌核发育草鱼群体的遗传多样性及雌核发育草鱼个体的基因纯合度进行分析。所得结果与结论如下： (1)在所选28对鲤鱼的微卫星引物中，经过反应条件的优化，筛选到了7对(25％)引物能在草鱼基因组中扩增出重复性好而且清晰的特异性条带，其中的4对(占14.3％)引物扩增的特异条带在群体中具有多态性。说明部分鲤鱼的微卫星引物对草鱼基因组分析具有适用性。 (2)7个微卫星位点上湘江野生草鱼群体的平均等位基因数Na、平均观测杂合度H_o、平均期望杂合度H_e、有效等位基因数N_e分别为2、0.3571、0.2992、1.6791，遗传相似性为0.8043。提示湘江野生草鱼群体的遗传多样性比较匮乏。 (3)两个雌核发育群体的13尾草鱼个体，在5个微卫星位点上的等位基因均达到纯合，占全部位点的71.4％；在1个位点上，部分个体基因纯合，部分个体基因杂合，占全部位点的14.3％。表明实验获得的雌核发育草鱼个体等位基因已有较高纯合程度。令人惊异的是在1个微卫星位点上，13尾草鱼个体均扩增出两个产物。推测这个位点在草鱼基因组中可能存在多拷贝。是否如此，尚有待进一步研究。 (4)雌核发育草鱼A群体在五个微卫星位点上达到完全的遗传同质性，群体内个体的基因型完全一致。雌核发育草鱼B群体在四个位点上达到完全的遗传同质性。雌核发育草鱼A、B两群体内的相似性系数分别为0.9091、0.8699。表明通过抑制第一次卵裂获得的两个雌核发育草鱼群体具有高遗传纯合度。 (5)两个雌核发育草鱼A、B群体与普通草鱼之间的遗传距离分别约是0.1620、0.1621，三个群体在7个微卫星位点上等位基因频率的分布无明显差异，提示两个雌核发育草鱼群体与野生群体之间无显著的遗传分化。更多还原

【Abstract】 Microsatellite primers which can be used for genomic analysis of grass carp were screened from 28 pairs of common carp microsatellite primers with methods of polymerase chain reaction (PCR) and polyacrylamide gel electrophoresis(PAGE). Then these primers were employed to investigate the genetic diversity of wild grass carp population of Xiang-jiang River and the mitogynogenetic grass carp groups induced by the heat-shocking of eggs.The main results and conclusions were as follows:(1) On the optimized conditions of PCR, total 7 out of 28 (about 25%) pairs of primers of common carp microsatellite loci were proved to be able to amplify successfully with model DNA of grass carp and showed clear and specific bands in PAG repeatedly. And 4 pairs of primers (about 14%) showed the polymorphic in population of the wild grass carp. These results indicated that some microsatellite primers developed from common carp could be applied to analyze the genome of grass carp.(2) The average allele numbers (N) were 2 in the wild grass carp population of Xiang-jiang River at 7 microsatellite loci. The average observation heterozygosity (H0) was 0.3571. The average expectation heterozygosity (He) was 0.2992.The effective allele number (Ne) was 1.6791. And the genetic similarity was 0.8043. These results indicated that the genetic diversity in wild grass carp population of Xiang-jiang River was poor.(3) At five out of seven (about 71.4%) microsatellite loci, all 13 individuals of grass carps of the two gynogentic populations (A and B) were homozygous. And at one locus (about 14.3%), some fish have showed homozygous and some heterozygous. These results showed that there was a high proportion of homozygous microsatellite loci in the gynogenetic grass carps. It was surprising that two PCR products wereobserved in all of the 13 individuals of two gynogenetic grass carp groups when their genomic DNA were amplified using the primer MFW1. It was presumed that there were two copies of this microsatellite sequence in the genome of the grass carp, which was needed to test in further studies.(4) Five microsatellite loci of the gynogenetic grass carp population A and four of the population B was genetically homogenous, meaning that each individual in the population had the same genotype at these loci. The genetic similarity index in the gynogenetic grass carp population A and B were 0.9091 and 0.8699 respectively. From these results a conclusion could be made that there was high genetic homozygosity in the two gynogenetic populations.(5) The genetic distance between the wild and gynogenetic grass carp population A was 0.1620 and that between the wild and gynogenetic grass carp population B was 0.1621. It seemed that there were some variations between the gynogenetic and wild populations. But the fact that no significant difference has been detected on the frequencies distribution of alleles of every microsatellite locus among the three populations by t-test indicated that the level of differentiation between the wild and gynogenetic grass carp populations was low since the variation was mainly caused by rare alleles. However, the lost of rare alleles in two populations of the gynogenetic grass carp should be considered in genetic breeding.更多还原