北太平洋柔鱼微卫星标记的筛选及遗传多样性

采用(AC)12、(AG)12两种生物素探针,通过磁珠富集法构建了柔鱼部分基因组微卫星富集文库。68个阳性克隆中有60个含有微卫星序列,重复次数在10次以上的占86.84%,最高重复次数为33次。其中,完美型微卫星占60.53%,非完美型微卫星占36.84%,混合型微卫星占2.63%。除探针使用的AC/TG、AG/TC重复外,还得到ACAG、AGAC重复序列。利用筛选出的8个微卫星位点对北太平洋柔鱼6个群体的遗传多样性及遗传结构进行分析。结果表明,8个微卫星位点均为高度多态性位点(PIC=0.787—0.987),位点Bo103与位点Bo105极显著偏离Hardy-Weinberg平衡(P0.01)。6个地理位置的柔鱼群体显示出较高的遗传多样性水平(Ho=0.672—0.761,He=0.808—0.851)。两两群体间的Fst值以及AMOVA分析结果均表明,群体间遗传分化不显著(Fst=0.00559,P0.05),遗传差异主要来自于个体间。基于Nei's遗传距离的UPGMA聚类树显示,北太平洋东北部2个柔鱼群体(NE1、NE3)聚为一类,西北部3个群体(NW1、NW2、NW3)与东北部1个群体(NE2)另聚为一类,且群体NW1与群体NE2亲缘关系最近,遗传距离与地理距离线性相关分析没有呈现出正相关性(R=0.175,P0.05)。遗传结构分析结果推断北太平洋柔鱼存在1个理论群。柔鱼个体具有较强的游泳能力,在海流的作用下,群体之间存在较强的基因交流。建议今后在柔鱼资源开发利用过程中将北太平洋柔鱼看作1个管理单元。

Isolation and genetic diversity of microsatellite DNA of Ommastrephes bartramii in the North Pcific Ocean

The neon flying squid Ommastrephes bartramii is found circumglobally in subtropical and temperate waters in the world and supports important fisheries in the North Pacific Ocean. The population of the North Pacific O. bartramii comprises a fall spawning cohort and a winter-spring spawning cohort. Most of the fall cohort occurs east of 170°E, whereas the winter-spring cohort is widely distributed across the North Pacific Ocean. Two spawning cohorts can be further subdivided into four stocks: a central and an eastern stock of the fall cohort separated near 160°W, and a western and a central-eastern stock of the winter-spring cohort separated near 170°E in which the fall cohort was the target of a driftnet fishery before 1993 while the west stock of winter-spring cohort became main fishing target of an international jigging fishery. Being a short-lived ecological opportunist, neon flying squid exhibits large fluctuations in stock abundance and distribution on the feeding grounds and large fluctuating recruitment on the spawning grounds in response to environmental variability. In order to improve our understanding of population structure, the genetic diversity of O. bartramii populations in the North Pacific Ocean was examined using microsatellite markers. Two types of biotin-labeled oligos were employed and part of microsatellite-enhanced genomic library of O. bartramii was constructed by the method of streptavidin magnetic beads. Sixty of 68 positive clones comprised microsatellite sequences and repeat number of more than 10 accounted for 86.84% with the largest one of 33. Of these were 60.53% perfect, 36.84% were imperfect, and the rest were compound type (2.63%). Repeats including (ACAG)_n and (AGAC)_n were also detected except that of AC/TG and AG/TC. Microsatellite markers were isolated and screened, and 8 of them were then used for the analysis of genetic diversity and genetic structure of six populations of O. bartramii in the North Pcific Ocean. Results showed that all the 8 loci were highly polymorphic (PIC=0.787-0.987), and locus Bo103 and locus Bo105 were extremely significant deviated from Hardy-Weinberg equilibrium (P < 0.01). All of the 6 different geographic populations showed high genetic diversities (H_o=0.672-0.761, H_e=0.808-0.851). Pairwise F test and AMOVA analysis detected no significant genetic differentiations among populations, and genetic differences mainly came from individuals. UPGMA clustering tree based on Nei''s genetic distance demonstrated that two stocks (NE1 and NE3) in the northeast of the North Pcific Ocean clustered together, while the three stocks (NW1, NW2 and NW3) in the western part as well as one stock (NE2) in the eastern part clustered together, and the genetic distance between NW1 and NE2 was closer. Relations between genetic distance and geographic distance did not show positive correlation (R=0.175, P > 0.05), indicating that different populations of O. Bartramii could mingle together. Genetic structure simulation analyses suggested that there was only one logic population of all individuals, and extensive gene exchange might occur among the populations, which could be attributed to the influence of ocean currents and swimming ability of O. bartramii individuals. The North Pacific O. bartramii should be considered as a single stock in their assessment and management.