中华鲟生长激素基因的cDNA序列结构与进化分析

采用RT—PCR方法从中华鲟脑垂体RNA中克隆出645bp的生长激素（growth hormone,GH）eDNA全序列,推导出的GH由214个氨基酸组成,其中前24个氨基酸为信号肽部分。与其它脊椎动物的37个极端保守的氨基酸残基相比,中华鲟有1个氨基酸的差异,即152位上的天冬氨酸变为谷氨酸。中华鲟GH氨基酸保守区可分为5个区域,GD1～GD4区域位于分子外侧,为GH与受体结合区域,与GH活性有关,GD5位于分子内部,与GH分子结构和稳定性有关。中华鲟与俄罗斯鲟的GH序列遗传距离最小,为0．012;与弓鳍鱼、4种哺乳动物和欧洲鳗鲡的遗传距离分别为0．202、0．249～0．260和0．370;而与其它11种硬骨鱼类的遗传距离较大,为0．514～0．601。NJ系统进化树结果显示,中华鲟、俄罗斯鲟、弓鳍鱼、4种哺乳动物和欧洲鳗鲡聚成一支,而其它11种硬骨鱼类聚为另一支。以上结果表明,中华鲟GH与俄罗斯鲟、弓鳍鱼、哺乳动物和欧洲鳗鲡的GH更为相似,而与其它硬骨鱼类的GH同源性较低。     

大量生产鳗鲡生长激素

日本东京协和发酵株式会社和东京大学海洋研究所共同研究出一种利用生物工程生产鳗鲡生长激素的新技术，监且在东京召开的日本分子生物学会上发表了论文。     

草鱼生长激素cDNA基因的克隆及序列分析

通过反转录PCR技术，从草鱼脑垂体的总RNA中获得一特异cDNA片段，经克隆后测序，结果表明，该片段长633bp，与报道的草鱼生长激素基因相比，有10个碱基的差异，而推导的蛋白质仅1个氨基酸残基的差异，推测为草鱼生长激素cDNA基因。     

鲮生长激素基因cDNA的分子克隆和序列分析

根据生长激素(growth hormone,GH)基因的保守性序列设计1对引物,利用RT—PCR技术从鲮(Cirrhina molitorella)脑垂体组织中克隆出经生长激素成熟肽cDNA片段,将纯化的DNA片段克隆到pUCm—T载体上。克隆的鲮GH cDNA开放阅读框全长567bp,编码由188氨基酸残基组成的生长激素成熟肽。序列分析结果表明,鲮GH成熟肽氨基酸序列与另2种鲮—印鲮(Cirrhina mrigal)和南亚野鲮(Labeo rohita)的成熟肽氨基酸序列同源性分别为87％和83％,与鲤的成熟肽氨基酸序列同源性为96％,并对9个科的17种鱼进行了分子系统进化树分析,结果与根据传统的形态学和生化特征分类进化地位基本一致。本研究所克隆的基因序列和推测的蛋白质序列已登录入GeoBank、EMBL和DDBJ基因库,序列号分别为AF458105和从AAL151107．1。     

兰州鲇生长激素(GH)基因的克隆及序列分析

利用长片段PCR及T-A克隆技术,从兰州鲇(Silurus lanzhouensis)基因组DNA中首次克隆兰州鲇GH基因全序列并提交至Genbank获得登录号KM215221,同时,使用DNAstar等生物信息学软件进行序列分析研究。兰州鲇GH基因全长2 067 bp,由5个外显子和4个内含子组成,完整编码序列(Complete coding sequence,CDS)为603 bp。5个外显子大小分别为10、140、117、132和204 bp;4个内含子大小分别为229、103、565和103 bp;内含子与外显子的连接区序列遵循基因组成规则。编码区编码1条由200个氨基酸残基组成的蛋白质多肽。5个科8种鲶形目鱼类生长激素基因编码区序列同源性比较表明,它们之间GH基因编码区序列有较高的同源性,平均达到了92.4%,其中,兰州鲇和南方大口鲇之间的同源性最高,为99.5%,兰州鲇和革胡子鲇之间的同源性最低,为90.0%,并构建了系统发育进化树。同时也表明,GH基因编码区序列具有较高的保守性,系统发育研究中具有较高的参考价值。     

翘嘴鲌两种生长激素受体基因结构及微卫星多态性与生长性状的相关性

为了更好地研究翘嘴鲌两种生长激素受体(GHR)的结构和功能,实验以翘嘴鲌转录组中获得的mRNA为基础,对其DNA序列进行了克隆。在进行生物信息学分析的同时,对其中的多态性微卫星位点在120尾同批繁殖、同塘养殖的翘嘴鲌个体中进行了分析。GHR1的cDNA序列长度为3 498 bp,开放阅读框(ORF)为1 818 bp,编码605个氨基酸;GHR2的cDNA序列长度为1 743 bp,ORF为1 743 bp,编码580个氨基酸;GHR1和GHR2氨基酸序列均由信号肽、胞外区、跨膜区、胞内区组成,相似度为37.2%。二者在结构上存在较大的差异:GHR1胞外区有7个半胱氨酸残基,而GHR2只有5个,且GHR1比GHR2多3个N-糖基化位点;在胞内区,GHR1存在10个酪氨酸残基而GHR2只有5个,这些差异表明二者可能具有不同的生物学功能。同源氨基酸序列比对发现,GHR与其他鲤科鱼类的同源基因保守性较高。翘嘴鲌2个GHR各包含9个内含子,其中GHR1内含子1和2序列在10 kb以上,本实验没有对其进行扩增。所获得的序列中共发现了6个微卫星位点:GHR1中微卫星位点(CT)₆     

齐口裂腹鱼生长激素基因全长cDNA克隆与序列分析

鱼类生长激素是一种单一肽链蛋白激素,主要作用是促进机体的生长.采用RT-PCR方法,首次从齐口裂腹鱼(Schizothorax prenanti)脑垂体总RNA中克隆出生长激素(gFowth hormone,GH)cDNA的开放阅读框ORF(open reading frame)序列,全长633bp.经DNA Star软件分析,该基因编码210个氨基酸,分子量为23.55 kD,等电点为5.46,含有1个疏水区和1个N-糖基化位点.空间结构含有4个α螺旋和4个β折叠,形成2个二硫键.序列比较发现,齐口裂腹鱼GH与草鱼、鲤的GH同源性分别为99.8%、99.4%;与黄鳝、花鲈和大黄鱼的GH同源性分别为57.4%、63.5%和64.2%.     

全鱼生长激素基因构建的意义

<正> 随着人们对高蛋白食品要求的增加,可以预期水产养殖业将逐渐承担起满足人类需求的重任。因而培育个体大、生长迅速的鱼类优良品种,其经济价值的潜力是显而易见的。现代分子生物学的迅速发展,使体外进行DNA操作已成为现实。转基因鱼的出现就是分子生物学技术在水产研究方面的应用实例。目前,转基因鱼的研究,主要集中在鱼类加速生长、抗病性及抗低温能力提高等方面。已有的成果表明,这是创造鱼类新品种的有效途径。     

澳洲鳗鲡微卫星分子标记的筛选与检测

采用小片段克隆法构建了澳洲鳗鲡(Anguilla australis)的部分基因组文库,并用地高辛标记的(CA)15作探针筛选阳性克隆,共获得76条微卫星序列.其中10次以下CA连续重复序列占83.67%,没有检测到30次以上的连续重复序列.根据微卫星序列两端足够长的侧翼序列,设计引物55对,选择合成引物26对,用澳洲鳗鲡3个个体的混合基因组进行引物筛选,其中的18对具有清晰的扩增条带.将筛选出的18对引物对澳洲鳗鲡1个群体的40个个体进行了遗传多样性分析,其中1对引物扩增产物为单态,17对扩增产物呈现多态;17对扩增多态的引物在40个个体中扩增出等位基因数目为5～14,平均为9个.该澳洲鳗鲡群体的PIC、Ho、He的平均值分别为0.715 7、0.677 9、0.7374,所有位点均符合哈迪-温伯格平衡(P=0.05),结果证明这17个微卫星位点适于澳洲鳗鲡群体结构的研究分析.[中国水产科学,2009,16(1):133-138]     

转生长激素基因鲤鱼的生长研究

本报导了生长激素基因对鲤鱼生长的影响。经池塘对照实验表明，转生长激素基因鲤鱼的快速生长效应比正常鲤鱼明显，而且由转生长激素基因阳性性成熟鲤鱼经人工催产繁殖的子一代群体中，仍表现出快速生长的特性。这一结果，为能获得一个转外源生长激素基因并具快速生长特性的遗传稳定的转基固鲤鱼核心群体提供了可靠的数据。     

用RAPD技术研究3种鳗鱼的种质鉴定

利用RAPD技术从32个随机引物中筛选出20个引物,对日本鳗(Anguillajaponicus)、欧洲鳗(Anguillaanguilla)和美洲鳗(Anguillarostrata)的DNA进行扩增,均得到1～13条大小不等的DNA片段,长度大部分在500～2300bp,其中14个引物共扩增出37个特异性标记,可用来区分这3种鳗鱼。日本鳗和欧洲鳗和美洲鳗种群内相似系数分别为0.885,0.717,0.686;种间遗传距离分别为:日本鳗和欧洲鳗0.747,日本鳗和美洲鳗0.668,欧洲鳗和美洲鳗0.211。这与形态学分析的结果相一致。     

Age and growth of yellow eel, Anguilla anguilla (L.), determined by two different methods

Abstract– Ageing and calculation of growth rate of eels has always been controversial, and remarkably little validation has been reported. This article describes the growth of yellow eels from three lakes in the Burrishoole system, Ireland, as determined by back-calculation from otoliths prepared by burning and cracking. Eels captured by fyke net ranged in length from 25.0 cm to 98.5 cm and the ages ranged from 6 to 49 years. Back-calculated growth was slow, with mean annual increments of 2.08 cm, 1.40 cm and 1.97 cm for the three lakes. Yellow eels captured in the Burrishoole system in 1987 and 1988 were marked by Pan Jet inoculated alcian blue dye and released back into the three lakes. Recaptures made over the following three years yielded considerable growth data. Mean annual growth rates recorded were 2.30 cm, 1.07 cm and 0.97 cm. These data compare favorably with the calculated growth rates, thereby validating the otolith analysis for the eels in the Burrishoole system.     

养殖鳗鲡红头病的研究

1995年2月—1996年10月,对欧洲鳗鲡红头病进行了病原菌分离、人工感染、病原菌生理、生化特性和药物敏感性的测定,观察了病理组织变化。结果认为病原菌鲁氏不动杆菌为革兰氏染色阴性、无动力的双杆状菌,接触酶阳性,氧化酶、旷V—P反应、MR反应、吲哚反应、H_2S产生阴性、对四环素、强力霉素、妥布霉素敏感。主要病变为头部水肿、出血、鳃丝水肿、溃烂,肝脏细胞实质性坏死,形成坏死结疖、肾小管上皮层分解,血细胞浸润形成溃疡灶、脾脏充血、肿大、细胞相互融合。     

Variation in numbers of eel Anguilla anguilla caught by constant effort in an Irish lake, 1981–1994

Abstract– A total of 8,903 eels were collected in 503 fyke-net samples during the months March to December from 1981 to 1994 in a small bay in Lough Derg, a mesotrophic lake. Variations in catch between months and between years are described. In addition to expected decreases in catch in the colder months, a regular reduction was observed in June. Low catches were observed from 1985 to 1987, coinciding with low water temperatures. A very low catch in August 1994 may have been a chance variation but could have been a first indication of the effects of poor recruitment or of increased commercial fishing effort. The results support the initial hypothesis that a long-term study is required to determine the extent of variation in an eel population.     

Movements of the European eel (Anguilla anguilla) in a chalk stream

Abstract – The movements of wild European eels (Anguilla anguilla L.) were monitored continuously over a 2‐year period in a tributary of the River Itchen, Hampshire, UK, using a passive integrated transponder (PIT) antenna system. The time of these movements was then related to a number of monitored and calculated environmental parameters. No eel movements were recorded in the winter months when mean stream temperature was below 10 °C. The numbers of movements peaked in May, when mean water temperatures and day length were increasing. The movement of eels was significantly (P < 0.001, R² = 0.45, N = 72) correlated with the time of sunset, with the majority (72%) recorded during the hours of darkness. The results of this investigation have implications for the design/timing of surveys of yellow eel populations and their habitat associations and suggest that successfully determining the microhabitat preferences of active resident eels will require remote monitoring throughout the 24‐h period.     

Effects of improving longitudinal connectivity on colonisation and distribution of European eel in the Loire catchment, France

Abstract –  Major migration barriers have been identified as an important factor in the drastic reduction of freshwater eel stocks at the catchment scale. In this study, we developed a way to assist decision making in the management of migration barriers so as to increase eel presence throughout the Loire catchment (France). Models were developed according to eel size-class and various different management strategies to improve accessibility, taking into account: (i) all the migration barriers from the tidal limit to different upstream limits (downstream–upstream strategy), (ii) all the migration barriers with a given degree of passability (migration barrier passability class strategy), (iii) all the migration barriers in a given sub-catchment area (sub-catchment strategy) and (iv) all the migration barriers in the entire Loire catchment (global strategy). Eel were actually found to occur in the Loire at about 50% of the sites sampled. The improvement of habitat accessibility by removing all obstacles to colonisation or installing fish passes, for example (global strategy), would lead to eels being found throughout the whole river. If this management strategy is impossible because the high cost, two others, also producing significant eel continental colonisation must be implemented. Either all barriers from the tidal limit with medium passability and/or migration barriers in the sub-catchment area which is closest to the sea and least accessible must be managed. In both cases, the area significantly exploited by eels and the probability that this species will be present will be doubled. However, these results can only be counted on if the fluvial recruitment increases significantly.     

恩诺沙星及其代谢物环丙沙星在欧洲鳗鲡体内的代谢动力学

采用高效液相色谱法,研究药饵口灌给药途径下,恩诺沙星及其代谢物环丙沙星在欧洲鳗鲡(Anguilla anguilla)体内的药代动力学。欧洲鳗鲡口灌给药恩诺沙星10 mg/kg后,其血浆、肌肉和肝脏中药物时量曲线关系符合一级吸收的二室开放动力学模型。恩诺沙星在欧洲鳗鲡不同组织中分布较广,血浆、肌肉和肝脏的Vd分别为6.362 L/kg、8.081 L/kg和15.870 L/kg;恩诺沙星在鳗鲡体内消除较慢,在血浆、肌肉和肝脏中的消除半衰期(tβ1/2)分别为161.10 h、333.21 h和611.26 h,总体清除率(CLs)分别为27.4 mL/(kg.h)、16.8 g/(kg.h)和18.0 g/(kg.h)。代谢物环丙沙星在鳗鲡血浆、肌肉和肝脏中药物水平的变化趋势与恩诺沙星基本相似,呈现出多峰现象,但3种组织中环丙沙星出现第一个药峰时间分别为给药后第24小时、24小时和12小时,3种组织中环丙沙星峰值水平肝脏中最高、肌肉中次之、血浆中最低,环丙沙星在肌肉和肝脏中的消除速率比较缓慢。鉴于恩诺沙星和其代谢物环丙沙星在欧洲鳗鲡体内消除较慢,建议养成阶段使用其他药物。     

Implications of the precautionary approach for the management of the European eel, Anguilla anguilla

The European eel, Anguilla anguilla (L.), has a complex life history and many aspects of the biology and population dynamics of this species remain unknown or, at best, poorly understood. Relatively little is also known about the status of the stocks and fisheries, but available data suggest that recruitment of glass eels has been falling for the last 20 years and is at historically low levels. Yellow and silver eel catches have also been falling in many parts of the species range over a similar time‐scale. Re‐examination of the principles applied to fisheries management over recent years has resulted in the adoption of a ‘precautionary approach’ to the conservation, management and exploitation of fish stocks, and in an explicit need to take account of uncertainties in management to reduce risks to stocks and their environment. Such an approach is highly relevant to the management of the European eel and requires that urgent consideration is given to harvest strategies and decision structures for the national and international management of stocks and fisheries. Provisional biological reference levels should be established to provide an equable assessment of the status of stocks in all parts of Europe and to evaluate the need for management measures in all fisheries. These will need to be reviewed as further information comes available. Monitoring and research on eel stocks should therefore be enhanced and co‐ordinated to improve our understanding of the status of stocks throughout Europe and the biology of the species.     

Decreasing eel stocks: survival of the fattest?

Abstract –  Since the 1980s, the European eel Anguilla anguilla stock is in steep decline. Lipid reserves are essential to cover energetic requirements for silver eel migration and reproduction. Two large and independent data sets from Belgium and The Netherlands show an average one-third decrease in fat contents of yellow eels over the past 15 years. Also Le Cren's relative condition factor decreased. On the basis of the somatic energy reserves, reproductive potential of eels from various latitudes over Europe was estimated, assuming fat levels in yellow eel are indicative of those in silver eels. Only large individuals, females as well as males, with high lipid content seem to be able to contribute to the spawning stock. The decrease in fat content in yellow eels may be a key element in the stock decline and raises serious concerns about the chances of the stock to recover.     

Spatial organisation of European eel (Anguilla anguilla L.) in a small catchment

Abstract –  Modelling-governing patterns of European eel ( Anguilla anguilla L.) distribution of four eel size classes (<150, 150–300, 300–450 and >450 mm) in the Frémur basin (northwest France) was done using artificial neural network (ANN) techniques and ecological profiles. Our results demonstrate the high predictive power of the ANN models. Some macro- and microscale factors, such as distance from the sea, depth and flow velocity, have the most significant influence on the models. Influence of distance from the sea appears to be very different from the spatial organisation usually described in river systems. In fact, the general tendencies of total eel densities according to the distance from the sea showed that densities increase weakly upstream. Another outcome was the variations in habitat preference according to the eel size, even if this species is spread over practically every type of microhabitat. Small eels were mainly found in shallow habitats with strong abundance of aquatic vegetation, whereas large eels tend to be found in intermediate to high depth with small to intermediate abundance of aquatic vegetation. Finally, we hypothesise that European eels change behaviour and microhabitat characteristic preference around a size of 300 mm.