黄姑鱼雌核发育诱导及鉴定  

Induction and identification of gynogenesis in Nibea albiflora

作　　者：杨育凯[1] 谢仰杰[1] 蔡明夷[1] 简林江[1] 陈庆凯 楼宝[3] 王志勇[1] 

机构地区：[1]集美大学水产学院农业部东海海水健康养殖重点实验室,福建厦门361021 [2]福建省宁德市水产技术推广站,福建宁德352100 [3]浙江省海洋水产研究所,浙江舟山316100

出　　处：《水产学报》2013年第9期1297-1303,共7页Journal of Fisheries of China

基　　金：国家科技基础条件平台资助;公益性行业(农业)科研专项(200903046-05);集美大学创新团队科研基金(2010A002);福建省教育厅A类科技项目(JA12189);浙江省重大科技专项(2012C12907-8)

摘　　要：为建立黄姑鱼雌核发育诱导方法,实验利用紫外线照射使黄姑鱼精子遗传失活,与卵子授精后再通过冷休克抑制极体排放进行倍性恢复,成功诱导出黄姑鱼雌核发育二倍体。精子经强度为3 800μW/(cm2.s)的紫外线照射10～100 s,受精卵孵化率呈现明显的Hertwig效应;当照射时长达到60 s以上,各实验组全部孵出仔鱼均呈现单倍体综合症。精子的紫外线照射时间、受精卵冷休克起始和持续时间等三因素三水平正交实验结果表明,精子经紫外线照射60 s,受精2 min后卵子在3～4℃海水中持续冷休克10 min为最佳诱导条件组合,可以获得最高的孵化率(16%)。最佳组合仔鱼形态和细胞相对DNA含量与正常二倍体一致,经微卫星标记检验证明全部不含有父本基因,为雌核发育二倍体。实验报道了成功诱导黄姑鱼雌核发育二倍体的条件,为进一步开展黄姑鱼良种选育和性别控制工作奠定了基础。In order to establish a procedure to induce artificial gynogenesis for genetic improvement and sex control studies in Nibea albiflora,ultraviolet irradiation was used to genetically inactivate the sperm and cold shock was used to induce the chromosome duplication of the eggs.Relative DNA content in newly-hatched larvae of candidate haploid gynogens,candidate diploid gynogens and the control groups from normal fetilization were mensurated,and paternity test was made using SSR markers for confirmation of gynogenesis.The gynogenetic diploid was successfully induced by activating egg development with UV-irradiated sperm combined with cold shock to prevent extrusion of the second polar body.UV irradiation time range was preliminarily determined by an insemination trial with the sperms irradiated by different doses of UV.The fertilized egg hatchability exhibited typical Hertwig effect when sperm irradiation time was 0-100 seconds at a UV intensity of 3 800 μW/（cm2·s）.All newly-hatched larvae exhibited typical haploid syndrome when the UV irradiation time was over 60 seconds.In addition,we designed an orthogonal experiment to further optimize UV irradiation time,cold shock initiation time and cold shock duration time.The orthogonal experiment results showed that the hatching rate was highest（16%）in the following conditions：sperm irradiation time was 60 seconds,cold shock temperature was 3-4 ℃,cold shock initiation time was 2 minutes after fertilization and cold shock duration time was 10 minutes.The larvae of the optimal group have the same morphological characteristics and cellular DNA content with normal diploids.A further paternity test with five compatible SSR primers of Larimichthys crocea proved that the larvae in the optimal group were all gynogenetic diploids without paternal specific alleles.

关 键 词：黄姑鱼 雌核发育 冷休克 DNA相对含量 微卫星标记 

分 类 号：Q343.3[生物学—遗传学] S917.4[农业科学—水产科学]