基于GBS技术的半滑舌鳎野生与养殖群体遗传多样性分析

目前半滑舌鳎的人工繁育中普遍存在近亲繁殖现象,种质资源和优良性状无法保障,关于半滑舌鳎野生群体和养殖群体的群体结构、混杂程度、遗传多样性分析等研究工作近10a内没有持续跟进,随着大量的半滑舌鳎人工繁育苗种增殖放流入海,野生和养殖种群的混杂程度有待评估。本研究对半滑舌鳎3个海捕野生群体和3个养殖群体进行遗传多样性分析和种质资源状况研究,通过GBS技术对87个半滑舌鳎样本进行测序,产生的高质量序列数据量为55.12Gb,共获563109个高质量的SNP位点用于遗传多样性分析。通过SNP分析遗传多样性发现,野生群体的遗传多样性略高于养殖群体,个别养殖群体高于野生群体,说明近年来增殖放流对野生群体的遗传多样性有所影响,遗传距离最远的是天津野生群体和河北养殖群体。在种群分化方面, 6个群体间不存在明显分化,尚未形成明显的地理隔离,不过聚类分析还是可以把野生群体和养殖群体分成两个亚群。本研究发现了一些高分化位点,计划通过设计引物扩增该段序列找出能够区分野生和养殖群体的候选标记。     

半滑舌鳎雌鱼基因组Fosmid文库构建及分析

半滑舌鳎(Cynoglossus semilaevis)是我国的大型经济鱼类,其养殖发展迅速。有关半滑舌鳎的基因组文库构建和分析等方面的研究,目前尚未见报道。用流式细胞仪检测雌性半滑舌鳎基因组大小。从半滑舌鳎雌鱼肌肉提取基因组DNA,选取大小合适的DNA片段,经末端修复并回收后,再以Fosmid作为载体,构建半滑舌鳎雌鱼基因组文库。经检测,雌性半滑舌鳎基因组大小约为606.36 Mb。所构建的Fosmid文库含有49 920个克隆,重组率为96.88%;插入片段长度分布在33~45 kb之间,平均为39.2 kb;覆盖雌性半滑舌鳎基因组3.12倍;从文库中筛选得到单拷贝DNA序列的概率为95.6%。对培养第1天和第6天的克隆进行比较,结果表明文库稳定性高,培养过程中没有发生变化。     

半滑舌鳎的一种细胞凋亡抑制因子的克隆、鉴定与分析

克隆半滑舌鳎(Cynoglossus semilaevis)的1种细胞凋亡抑制因子survivin基因,并对其序列及表达模式进行分析.结果表明,该基因cDNA 序列全长704 bp(GenBank登录号为EU499314),包括86 bp5'非翻译区,444 bp开放阅读框以及含 poly(A)信号的174 bp的3'非翻译区,编码147个氨基酸.氨基酸序列同源性分析表明,半滑舌鳎survivin氨基酸序列与斑马鱼、非洲爪蟾、鸡、小鼠和人survivin氨基酸序列的同源性分别达到了63.3%,49.0%,43.7%,43.6%和41.5%.RT-PCR分析表明,survivin基因在半滑舌鳎成体雌鱼卵巢中大量表达,在雌鱼的肾脏中微量表达,在其它雌鱼的组织中无表达,在雄性的任何组织中均无表达.该基因在受精卵、多细胞和囊胚3个胚胎发育的早期阶段,大量表达,而随着胚胎发育的进程,survivin基因的表达逐渐减少,在孵化前尾芽期胚胎中只有微量表达,在孵化后5 d的仔鱼中,几乎检测不到survivin基因的表达.表明该基因可能参与卵子形成和胚胎的早期发育.     

半滑舌鳎雌性特异fosmid克隆的筛选及FISH定位

通过对半滑舌鳎雌鱼fosmid文库进行有序混合,成功构建了两步-三维PCR筛选体系,为后续的基因筛选奠定了基础。通过对半滑舌鳎W染色体文库序列进行引物设计,并分别在雌、雄鱼基因组DNA中筛选,获得1对雌鱼特异性引物。使用该引物对fosmid文库进行筛选,得到1个雌鱼特异性克隆,并通过荧光原位杂交技术将克隆定位到了W性染色体上,证实了使用该体系进行文库筛选的有效性以及雌鱼引物的特异性,建立了半滑舌鳎遗传性别的分子及细胞遗传学鉴定方法,为后续半滑舌鳎基因组及性染色体的研究奠定了基础。     

半滑舌鳎三倍体诱导研究

采用冷休克抑制第二极体排出的方法诱导了半滑舌鳎三倍体,并探讨了三倍体诱导的最佳条件.通过流式细胞仪倍性检测和染色体观察验证各组三倍化率.结果显示:在受精后5 min开始,将受精卵放在4℃海水中进行冷休克处理,持续时间为20 min,三倍体诱导率最高为35％.通过对三倍体染色体进行分析,辨认其雌雄染色体型为ZWW/ZZZ,出现频率为1∶1,没有发现ZZW型雌鱼,暗示半滑舌鳎Z染色体和W染色体之间的交叉和互换受到严重抑制.     

渤海湾半滑舌鳎及焦氏舌鳎的鱼卵和仔稚鱼的形态

半滑舌鳎Cynoglossus semilaevis Günther和焦氏舌鳎C.joyneri Günther 均隶属于舌鳎科Cyonglossidae的舌鳎属Cynoglossus,系广泛分布在我国近海的底层鱼类。它们具有一定的经济价值。其中半滑舌鳎是该属中个体较大的种类,最大体长可达800毫米以上。素有名贵鱼类之称。惟产量不高。该类鱼多栖息在海湾水域,洄游距离短,活动范围小,是适合增养殖的对象之一。 关于这两种舌鳎的早期形态资料迄今尚未     

温度对半滑舌鳎的生长、生化组成和能量收支的影响

研究了不同温度(16,19,22,25,28和31 ℃)对半滑舌鳎(Cynoglossus semilaevis Günther)幼鱼的生长、体成分组成和能量收支的影响.结果表明,随温度的升高,半滑舌鳎幼鱼生长率总体呈现先升高后降低的趋势.其中,在19～25 ℃温度范围内,半滑舌鳎幼鱼的特定生长率相对较高,在16和28 ℃下则有所降低,而在31 ℃下,半滑舌鳎幼鱼特定生长率则显著降低.本研究表明半滑舌鳎幼鱼特定生长率与温度符合二次曲线模型.半滑舌鳎摄食量随温度升高而逐渐增大,但在31 ℃时显著减小,饵料转化率则随温度的升高而降低.鱼体脂肪与能值均随温度升高而降低,蛋白质含量受温度影响不显著.温度对半滑舌鳎的能量收支影响显著,其中,生长能和代谢能主导半滑舌鳎的能量分配,生长能占摄食能比例则随温度的升高而降低,呼吸能占摄食能比例则随温度的升高而升高.本研究表明,半滑舌鳎幼鱼(13～37 g)适宜生长温度范围为18～25 ℃,而不同温度导致的半滑舌鳎摄食量和能量收支的差异可能是温度影响其生长的主要生理生态学机制.     

雄性半滑舌鳎GH基因多态性与激素水平及生长性状相关性分析

以雄性半滑舌鳎(Cynoglossus semilaevis)为研究材料,分析了其GH基因外显子区域的多态性,同时分析了各基因型与其体重、体长、激素水平等的关联性。PCR-SSCP和测序结果表明,外显子4有1个突变位点382bp(c.T382C)为非同义突变(p.Phe106Leu)。单因素方差分析显示,382bp位点与体重、去内脏重和血浆T3水平显著相关(P0.05)。通过多重比较,CC基因型体重和去内脏重((234.58±8.11)g和(210.23±7.55)g)显著高于TT基因型((201.64±13.04)g和(180.21±12.13)g);CC基因型(3.01±0.07)ng·mL-1的T3水平极显著高于TT基因型(2.12±0.12)ng·mL-1。结果表明,半滑舌鳎中GH基因的突变可以影响其激素水平和生长性状,382bp位点可以用作分子标记,从遗传学角度辅助半滑舌鳎优良品种的分子选育。     

花鲈人工育苗技术的研究

利用人工培育的亲鱼所获得的受精卵 ,孵化出仔鱼 30 0万尾 ,在水温 18～ 2 0℃、微充气的培育条件下 ,使用海水小球藻、轮虫、卤虫无节幼体和不同规格的微颗粒配合饲料进行饲喂 ,经 58d培育出平均全长 2 .3cm的苗种 132万尾 ,育苗成活率高达 64.1% ,单位水体最高出苗量达 1.33万尾 / m3;另外 ,还有 1.3～ 1.9cm的苗种 68万尾     

促激素及其受体在雄性半滑舌鳎脑垂体和精巢中的定位

应用兔抗血清对抗鱼类和哺乳类促激素及其受体的多克隆抗体对雄性半滑舌鳎脑垂体和精巢进行了免疫组织化学定位.其结果显示GnRHR、LHR和GtH免疫活性定位在不同发育时期的腺垂体和精巢中.GtH免疫阳性细胞的数量和免疫强度随着精巢发育成熟而明显增强.同时还发现睾酮（T）和雄激素受体（AR）在半滑舌鳎精巢中的定位有别于其他鱼类,尤其是雄激素受体（AR）免疫活性在精原细胞中的数量很少.依据这个特点文中还讨论了其可能与精巢发育的关系,同时证明了半滑舌鳎精巢发育像其他硬骨鱼类一样受激素的调控.     

Microbial Biomass and Organic Nutrients in the Deep-Sea Sediments of the Central Indian Ocean Basin

In order to assess the impact of deep-sea mining on the in situ benthic life, we measured the microbial standing stock and concentration of organic nutrients in the deep-sea sediments of the Central Indian Ocean Basin in the Indian pioneer area. Sediments were collected using box core and grab samples during September 1996. The total bacterial numbers ranged from 10 10 -10 11 cells per g -1 dry weight sediment. There was a marginal decrease in the number of bacteria from surface to 30 cm depth, though the subsurface section registered a higher number than did the surface. The highest numbers were encountered at depths of 4-8 cm. The retrievable number of bacteria were two orders less in comparison with the direct total counts of bacteria. An almost homogeneous distribution of bacteria, total organic carbon, living biomass, and lipids throughout the depth of cores indicates active microbial and benthic processes in the deep sea sediments. On the other hand, a uniform distribution of total counts of bacteria, carbohydrates, and total organic carbon in all the cores indicates their stable nature and suggests that they can serve as useful parameters for long-term monitoring of the area after the benthic disturbance. Further studies on temporal variability in this region would not only verify the observed norms of distribution of these variables but would also help to understand restabilization processes after the simulated benthic disturbance.     

Assessing the Validity of Seismo-Acoustic Predictor Equations for Obtaining Seabed and Sub-Surface Sediment Physical Properties

The interdependence between the seismo-acoustic properties of a marine sediment and its geotechnical/physical parameters has been known for many years, and it has been postulated that this should allow the extraction of geotechnical information from seismic data. Though in the literature many correlations have been published for the surficial layer, there is a lack of information for greater sediment depths. In this article, a desktop study on a synthetic seafloor model illustrates how the application of published near-surface prediction equations to subsurface sediments (up to several tens of meters burial depth) can lead to spurious predictions. To test this further, acoustic and geotechnical properties were measured on a number of sediment core samples, some of which were subjected to loading in acoustically-equipped consolidation cells (oedometers) to simulate greater burial depth conditions. For low effective pressures (representing small burial depths extending to around 10 meters subsurface), the general applicability of established relationships was confirmed: the prediction of porosity, bulk density, and mean grain size from acoustic velocity and impedance appears generally possible for the investigated sedimentary environments. As effective pressure increases through, the observed relationships deviate more and more from the established ones for the near-surface area. For the samples tested in this study, in some instances increasing pressure even resulted in decreasing velocities. There are several possible explanations for this abnormal behavior, including the presence of gas, overconsolidation, or bimodal grain size distribution. The results indicate that an appropriate depth correction must be introduced into the published prediction equations in order to obtain reliable estimates of physical sediment properties for greater subsurface depths.     

A Review of Some Environmental Issues Affecting Marine Mining

This article reviews information recently available from existing marine and coastal mining for responses to environmental issues affecting marine mining at different depths. It is particularly but not exclusively concerned with those issues affecting seabed biodiversity impact and recovery. Much information has been gathered in the past 10 years from shallow mining operations for construction aggregate, diamonds, and gold, from coastal mines discharging tailings to shallow and deep water, and from experimental deep mining tests. The responses to issues identified are summarized in a series of eight tables intended to facilitate site-specific consideration. Since impacts can spread widely in the surface mixing layer SML, and can affect the biologically productive euphotic zone, the main issues considered arise from the depth of mining relative to the SML of the sea. Where mining is below the SML, the issue is whether it is environmentally better to bring the extraction products to the surface vessel for processing (and waste discharge), or to process the extraction products as much as possible on the seabed. Responses to the issues need to be sitespecific, and dependent on adequate preoperational environmental impact and recovery prediction. For deep tailings disposal from a surface vessel, there are four important environmental unknowns: (1) the possible growth of "marine snow" (bacterial flocs) utilizing the enormous quantities of fine tailings particles (hundreds or thousands of metric tons per day) as nuclei for growth, (2) the possibility that local keystone plankton and nekton species may migrate diurnally down to and beyond the depth of deep discharge and hence be subjected to tailings impact at depth, (3) the burrow-up capability of deep benthos and their ability to survive high rates of tailings deposition, and (4) the pattern and rate of dispersion of a tailings density current through the deep water column from discharge point to seabed. Actions to obtain relevant information in general and site-specifically are suggested.     

Increased Particle Fluxes at the INDEX Site Attributable to Simulated Benthic Disturbance

Particle fluxes were measured 7 m above the sea bottom during the predisturbance, disturbance, and postdisturbance periods by using time series sediment traps attached to seven deep-sea moorings deployed in the INDEX experiment site in the Central Indian Basin. The predisturbance particle fluxes varied between 22.3 to 55.1 mg m -2 day -1 . Increased and variable particle fluxes were recorded by the sediment traps during the disturbance period. The increase observed was 0.5 to 4 times more than the background predisturbance fluxes. The increases in particle fluxes (~4 times) recorded by the sediment trap located in the southwestern direction (DMS-1) were the greatest, which could be the result of preferential movement of resuspended particles generated during the deep-sea benthic disturbance along the general current direction prevailing in this area during the experimental period. Also, the traps located closer to the disturbance area recorded greater fluxes than did the traps far away, across the Deep Sea Sediment Resuspension System path. This variability in recorded particle fluxes by the traps around the disturbance area clearly indicates that physical characteristics such as grain size and density of the resuspended particles produced during the disturbance had an important effect on particle movement. The postdisturbance measurements during ~5 days showed a reduction in particle fluxes of ~50%, indicating rapid particle settlement.     

海上大直径钢管桩打桩过程中桩周土强度弱化研究

海上大直径钢管桩打桩过程中,桩周土体受到强烈扰动而发生强度弱化,掌握桩周土体强度弱化规律对于准确预测打桩过程、保证工程安全具有重要意义。为研究土体强度弱化规律,开展了环剪试验模拟打桩对桩周土体的扰动,测试土体强度随剪切速率的变化规律,建立了描述土体强度弱化规律的拟合公式,引入到打桩分析软件中。研究结果表明：土体的强度折减程度不仅与土体本身的性质有关还受到土体的埋深和剪切速率的影响,埋深越深土体强度折减程度越低,剪切速率越高土体强度折减越高,在打桩分析中可采用这里推荐的线性折减方法来模拟不同深度处土体强度的折减规律。     

Geoacoustic Inversion via Genetic Algorithm and Its Application to Manganese Sediment Identification

An acoustic inversion method using a wide-band signal and two near field receivers is proposed and applied to multiple layered seabed models including a manganese sediment. The inversion problem can be formulated into a probabilistic model comprised of signals, a forward model, and additive noise. The forward model simulates wide-band signals, such as chirp signals, and is chosen to be the source-waveletconvolution plane wave modeling method. The wavelet matching technique, using weighted least-squares fitting, estimates the sediment sound-speed and thickness on which determination of the possible numerical ranges for a priori uniform distribution is based. The genetic algorithm is applied to a global optimization problem to find a maximum a posteriori solution for determined a priori search space. Here the object function is defined by an L 2 norm of the difference between measured and modeled signals. Not only the marginal pdf but also its statistics are calculated by numerical evaluation of integrals using the samples selected during importance sampling process of the genetic algorithm.     

Determination of In Situ Sediment Shear Strength from Advanced Piston Corer Pullout Forces

The advanced piston corer (APC) has been used by the Ocean Drilling Program since 1985 for recovering soft sediments from the ocean floor. The pullout force measured on extracting the core barrel from the sediment is shown to correlate with the average shear strength of the sediment core measured in the ship's laboratory. A simple rule of thumb is derived relating the shear strength of the sediment to the pullout force. Multiple APC holes at individual sites allow the consistency of the pullout measurements to be assessed. The effects of different operational procedures during APC coring are also explored. Although generally applicable, the correlation between pullout force and laboratory measurements of shear strength breaks down for some APC holes, possibly because of the disturbance of some sediment types during the APC coring process. A better understanding of the physical process of APC coring, and its effect on the properties of the sediment both inside and immediately outside the core barrel, would indicate what confidence can be put on the measurement of pullout force as a way of evaluating the in situ shear strength of deep sea sediments.     

Interannual Sea Level Variations and Annual Tides in the Northwestern Pacific

Specific properties of the interannual sea level variations and annual tides in the Northwestern Pacific were studied. Several tide stations were monitored. The monthly mean sea level for the year of 1995 was analyzed at each tide station. A seismic event in 1995, some tectonic activity around the subject area, and the Kuroshio (the oceanic western boundary current) may possibly contaminate results which would have occurred from the astronomical annual tide alone.     

《Acta Oceanologica Sinica》Vol. 33(2014) contents

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