三疣梭子蟹(Portunus trituberculatus)I 型Crustin抗菌肽的基因克隆与真核重组表达

Crustin 是目前报道广泛存在于甲壳类中的一类具有广谱抗菌活性的抗菌肽, 主要由甲壳类血淋巴细胞合成。本研究采用RACE 技术和真核表达技术, 进行了三疣梭子蟹Crustin 基因的克隆和重组表达研究。结果表明, 在三疣梭子蟹血细胞Crustin 基因cDNA 全序列中, 开放阅读框编码110个氨基酸残基的Crustin 前体, Crustin 前体由N 端21 个氨基酸残基的信号肽、富含Cys 结构域和C端WAP 结构域三部分组成, 属于I 型Crustin, 构建的分泌型真核表达质粒pVT102U/α-crustin 转化酿酒酵母S78, 经RT-PCR 和SDS-PAGE 电泳验证, 表明重组Crustin 表达成功。     

三疣梭子蟹(Portunus trituberculatus)Ⅰ型Crustin抗菌肽的基因克隆与真核重组表达

Crustin是目前报道广泛存在于甲壳类中的一类具有广谱抗菌活性的抗菌肽,主要由甲壳类血淋巴细胞合成。本研究采用RACE技术和真核表达技术,进行了三疣梭子蟹Crustin基因的克隆和重组表达研究。结果表明,在三疣梭子蟹血细胞Crustin基因cDNA全序列中,开放阅读框编码110个氨基酸残基的Crustin前体,Crustin前体由N端21个氨基酸残基的信号肽、富含Cys结构域和C端WAP结构域三部分组成,属于I型Crustin,构建的分泌型真核表达质粒pVT102U/α-crustin转化酿酒酵母S78,经RT-PCR和SDS-PAGE电泳验证,表明重组Crustin表达成功。     

中国对虾素在大肠杆菌(Escherichia coli)中的重组表达

中国对虾素是从中国对虾血细胞中克隆得到的一种抗菌肽。为了进一步研究中国对虾素(CHP)的功能并为制备特异性抗体作准备，采用大肠杆菌表达外源蛋白的方法，进行了对虾素原核表达的研究。根据从中国对虾血细胞中克隆得到的对虾素基因设计特异性引物扩增中国对虾素成熟肽基因(CHP)，插入原核表达载体pGEX-4T-1中，在E．coli BL21表达融合蛋白GST-CHP．结果表明，不同表达菌株的融合蛋白表达量为30％—34％，同一菌株在诱导5h后能达到最高表达量。利用GST亲和层析纯化融合蛋白，将融合蛋白用凝血酶裂解以得到CHP，其分子量约为5．6kDa，N-端测序结果与期望的成熟对虾肽序列一致。用液体生长抑制方法检测活性，表明重组GST-CHP蛋白及CHP均表现出对大肠杆菌的抑菌活性。     

养殖期中国对虾抗菌肽的表达

利用半定量RT-PCR方法,根据中国对虾(Fenneropenaeus chinensis)抗菌肽的cDNA序列,设计一对特异引物,首次对中国对虾养殖期(PL11期仔虾～稚虾)抗菌肽的表达进行了研究。结果在各时期样本所提取的总RNA中,均扩增出一条长度约为200bp的特异性条带,表明中国对虾养殖期不同阶段均有抗菌肽的表达,提示中国对虾抗菌肽基因为组成型表达,并可能是中国对虾抵抗微生物感染的一个重要分子基础。     

青蛤(Cyclina sinensis)溶菌酶基因在鳗弧菌(Vibrio anguillarum)刺激下的表达

利用构建的SMART cDNA文库和高通量测序方法,得到了青蛤溶菌酶相关基因的全长,采用荧光定量RT-PCR方法分析了c型溶菌酶基因在鳗弧菌刺激下的表达变化。结果表明,青蛤有c型溶菌酶和i型溶菌酶基因,分别编码155和182个氨基酸,c型溶菌酶信号肽为21个氨基酸并具有典型的LYZ1结构域,i型溶菌酶信号肽为19个氨基酸,其结构域为Destabilase domain结构,用于识别和切断谷氨酰胺-甲酰胺与赖氨酸ε-氨基之间的肽键。荧光定量PCR结果显示,在鳗弧菌刺激后6—24h,青蛤血细胞中c型溶菌酶的表达量出现明显上调的趋势,与对照组有显著性差异(P0.05),说明c型溶菌酶基因在青蛤的免疫应答中起重要的作用。     

中国明对虾Rel/NF-κB家族基因在弧菌刺激下的表达

利用RT-PCR和RACE技术获得了中国明对虾Rel／NF-κB家族成员FcRel基因的部分cDNA片段,并研究了该基因在弧菌刺激条件下的转录表达变化。结果表明,该cDNA编码320个氨基酸,包含一个完整的RHD结构域和部分IPT-NFkappaB结构域。FcRel基因的RHD结构域与其它物种的RHD结构域的分子系统学分析表明,中国明对虾的FcRel与果蝇的Relish聚在一起,然后与疟蚊、烟草天蛾等其它节肢动物的Relish聚为一类。在弧菌刺激时,FcRel在血细胞和淋巴器官中的转录表达显示出不同的变化模式。在弧菌注射后3h和5h,血细胞中FcRel的转录表达出现明显下调;而弧菌注射引起淋巴器官中FcRel基因明显的上调表达。说明FcRel基因的表达与弧菌刺激密切相关,提示Relish可能参与了对虾抗菌反应的信号转导通路。     

大黄鱼(Larimichthys crocea)I型干扰素基因的特征与表达分析

采用RT-PCR和RACE-PCR技术克隆了大黄鱼(Larimichthys crocea)Ⅰ型干扰素基因全长cDNA序列及其基因组序列,并利用荧光定量PCR技术研究了该基因在大黄鱼不同组织中的表达谱,以及副溶血弧菌(Vibrio parahaemolyticus)、LPS、polyI:C刺激后大黄鱼脾脏、肝脏和头肾组织中该基因在转录水平的表达变化。结果表明,大黄鱼Ⅰ型IFN基因全长cDNA为878bp,开放阅读框558bp,编码185个氨基酸。推测的大黄鱼Ⅰ型IFN氨基酸序列N端含有20个氨基酸的信号肽,其后包含一个保守的IFabd结构域。大黄鱼Ⅰ型IFN基因包含5个外显子,4个内含子,在大多数组织和细胞中都有表达,其中在血细胞中表达量最高,肌肉中表达量最低。PolyI:C刺激可诱导大黄鱼Ⅰ型干扰素基因在脾脏、头肾和肝脏中的表达量显著上调;LPS刺激可诱导大黄鱼Ⅰ型干扰素基因在脾脏和肝脏中的表达量显著上调;灭活的副溶血弧菌可诱导大黄鱼Ⅰ型干扰素基因在头肾和肝脏中的表达量显著上调。     

银鲳(Pampus argenteus)胰岛素样生长因子-I基因的克隆和表达分析

胰岛素样生长因子-I(Insulin-like growth factor-I,IGF-I)是影响脊椎动物生长、发育及代谢的重要调控因子。本研究采用RT-PCR和RACE技术,克隆了银鲳(Pampus argenteus)肝脏IGF-IcDNA序列,应用半定量RT-PCR、Real-time q PCR和原位杂交的方法检测了IGF-I的组织表达特性、在肝脏中的生长表达特性和IGF-I基因在肝脏中的定位。序列分析表明,IGF-I cDNA序列全长836bp,其5′非编码区128bp、3′非编码区92bp,开放阅读框(open reading frame,ORF)605bp,由此推导IGF-I前体蛋白由201个氨基酸组成;前体肽由信号肽、成熟肽、E肽三部分组成,其中信号肽59个氨基酸,成熟肽68个氨基酸,E肽74个氨基酸;成熟肽由B、C、A、D四个区域组成,E肽分析表明,银鲳IGF-I属Ea-4型。同源性比较结果表明,银鲳与同目鲈形目鱼类的IGF-I编码序列同源性较高,为83.52%—91.40%;与哺乳类、鸟类和爬行类的同源性较低。半定量RT-PCR和Real-time q PCR组织特异性表达结果显示,IGF-I m RNA在肝脏组织中的表达量最高,显著高于其它组织,肾脏、心脏、肌肉、脑、鳃、小肠、卵巢次之,嗅球、脾脏和胃中表达较低;半定量RT-PCR和Real-time q PCR不同生长阶段表达结果显示,IGF-I m RNA在30—50g肝脏组织中表达量最高(P0.05);IGF-I m RNA在肝脏中的原位杂交定位结果显示,在肝脏细胞中均有表达,阳性信号主要位于细胞质中,靠近细胞边缘处信号较强。     

大黄鱼抗菌肽hepcidin在巴斯德毕赤酵母中的表达及其产物的抑菌活性

Hepcidin是一类具有独特性质的小分子抗菌肽,克隆得到大黄鱼(Pseudosciana crocea)的hepcidin基因(Lyc-Hepc),cDNA序列全长585个核苷酸,编码一个由85个氨基酸组成的蛋白,包含24个氨基酸的信号肽、35个氨基酸的前导肽和26个氨基酸的成熟肽,成熟肽具有8个保守的半胱氨酸.将Lyc-Hepc 183bp的编码前导肽和成熟肽的前体肽(25aa-85aa)基因片段克隆到毕赤酵母表达载体pPICZαA中,然后电击转化巴斯德毕赤酵母SMD1168,经高浓度Zeocin筛选获得多个单克隆.挑取4个克隆进行培养,提取酵母基因组进行PCR检测,结果表明目的基因片段已成功整合到酵母菌SMD1168基因组中,获得酵母工程菌株SMD1168/pP1CZα-Lyc-Hepc.SDS-PAGE和WesternBlot分析表明酵母工程菌株在0.5％甲醇诱导培养72h后,Lyc-Hepc在培养上清中得到了分泌型表达.体外抑菌实验表明重组Lyc-Hepc蛋白能够抑制2株水产养殖病害菌嗜水气单胞菌和副溶血弧菌的生长.研究结果表明大黄鱼hepcidin可能在抗细菌免疫中发挥了作用.     

脊尾白虾(Exopalaemon carinicauda)组织蛋白酶D基因的克隆及其表达分析

采用RACE技术获得了总长为1853bp的脊尾白虾(Exopalaemon carinicauda)组织蛋白酶D基因全长cDNA序列。该基因5′和3′非编码区分别为120bp和572bp,开放阅读框为1161bp,推测编码386个氨基酸,预测分子量为42.36kDa,理论等电点为7.54,命名为EcCatD基因。同源性和系统进化分析表明,EcCatD基因与斑节对虾、美洲螯龙虾的相似性分别为86%和80%,与斑节对虾和美洲螯龙虾紧密聚为一支。荧光定量RT-PCR结果表明,EcCatD基因在血细胞中的相对表达量最高,其次为肝胰腺。该基因在鳗弧菌和WSSV感染后的脊尾白虾血细胞和肝胰腺中的表达量显著增加,并具有不同的时空表达趋势。结果表明EcCatD基因在脊尾白虾免疫反应中具有重要作用。     

The Gulf of California: Review of ecosystem status and sustainability challenges

The Gulf of California is unique because of its geographical location and conformation. It hosts diverse ecosystems and important fisheries that support industry and provide livelihood to coastal settlements. It is also the site of interests and problems, and an intense interaction among managers, producers, and conservationists. In this report, we scrutinize the abiotic (hydrography, climate, ocean circulation, and chemistry) and biotic (phyto- and zooplankton, fish, invertebrates, marine mammals, birds, and turtles) components of the marine ecosystem, and some particular aspects of climate variability, endemisms, harmful algal blooms, oxygen minimum layer, and pollution. We also review the current conditions and conflicts around the main fisheries (shrimp, small and large pelagic fishes, squid, artisanal and sportfishing), the most important human activity in the Gulf of California. We cover some aspects of management and conservation of fisheries, especially the claimed overexploitation of fish resources and the ecosystems, and review proposals for creating networks of marine protected areas. We conclude by identifying main needs for information and research, particularly the integration of data bases, the implementation of models and paleoreconstructions, establishment of monitoring programs, and the evaluation of fishing impacts and management actions.     

海洋生物多样性中心形成与演变机制的比较基因组研究进展

在大气圈、水圈、岩石圈和生物圈等多圈层相互作用下,全球范围内海洋生物多样性中心的地理位置、物种组成结构和优势度都呈动态变化.迄今,海洋生物多样性中心的形成与演变机制在生态、遗传和分子等层面正逐步得到解析.在生态层面,学界涌现出了物种形成中心、物种汇聚中心、物种重叠中心和物种保存中心等重要科学假说,解析了生物多样性分布格...     

红纤维虾形草(Phyllospadix iwatensis)种子形态及内部特征

海草床可以为海洋动物提供产卵、索饵、繁育等场所。目前, 海草床面积急剧下降, 其中我国关于虾形草属(Phyllospadix)海草的研究相对较少, 对于虾形草主要繁育载体种子的描述、显微观察和萌发过程方面存在较多空白。本研究以红纤维虾形草(Phyllospadix iwatensis)为研究对象, 对其生长环境、种子外部特征、内部结构、萌发过程等进行了定性定量描述。结果表明, 虾形草果实种子对于海洋环境有着外部形态与内部结构的适应性, 外部形态特点包括独特的果序和果实形状, 以及翅状结构和内侧鬃毛, 其中果序呈弯虾形, 平均长度3.4cm, 果实呈马掌形, 直径3~4mm, 内部结构特征包括胚乳占比大、果皮种皮解剖结构复杂等, 具有作为海岸带植被修复工具种的优势。本研究补充了目前对虾形草等海洋沉水植物种子的研究资料, 为虾形草海草种质库保护与海草床修复提供了建议和参考。     

Trade secrets: Renaming and mislabeling of seafood

As the global trade and market for seafood has grown, so have the twin problems of renaming and mislabeling. Resource scarcity, the potential for greater profits, and weak legislation have all encouraged incorrect labeling, the results of which include consumer losses, the subversion of eco-marketing, further degradation of fisheries resources, and even adverse effects on human health. This paper examines the extent and consequences of renaming and mislabeling seafood, the state of current legislation, and the importance of future policies, with particular attention to the US, where 80% of the seafood is imported and more than one-third of all fish are mislabeled. Policy recommendations include governments’ support for a global mandate to label species, country of origin, and catching or production method on all seafood with high penalties for infractions. Chain of custody standards, such as those recently implemented by the Marine Stewardship Council (MSC), should also be considered for adoption worldwide. To garner support for this legislation, consumers must become better acquainted and concerned with their seafood and its origins.     

养殖文蛤体内寄生的一种吸虫幼虫及宿主组织病理学

作为文蛤吸虫病害研究的基础资料,报道了寄生在文蛤体内的1种复殖吸虫幼虫的形态结构及宿主由此所产生的组织病理学变化。研究表明：该吸虫分别为咆蚴和尾蚴。咆蚴圆筒形,内含不同发育阶段的尾蚴。成熟尾蚴体成扁平舌状,具有口吸盘、腹吸盘,1对眼点和1条细长无刺的尾巴。该吸虫主要侵占生殖腺,少部分幼虫进入附近的消化盲囊、鳃、肾等部位。轻微感染者,生殖滤泡萎缩,生殖细胞发育滞后;严重感染的宿主,生殖腺完全被侵占并耗尽。消化盲囊、鳃、肾等器官组织也因幼虫感染呈现不同程度的病理学变化;严重者上皮细胞水肿或脱落,肌肉组织结构紊乱或溶解等。本工作同时显示,吸虫寄生可诱发宿主血淋巴细胞大量增生,并产生凝集、组织浸润及形成包囊等免疫反应。     

Estimation of Sediment Properties during Benthic Impact Experiments

Sediment properties, such as water content and density, have been used to estimate the dry and wet weights, as well as the volume of sediment recovered and discharged, during benthic impact experiments conducted in the Pacific and Indian Oceans. The estimates show that the weights of dry (355-1332 t) and wet sediment (1651-4888 t) recovered during the experiments, are not only a function of the total duration and distance covered during the experiment, but also depend on water content and density of the sediment. Estimation of dry sediment and its volumetric ratio in the discharge, are key parameters for calculating the discharged volume. The estimated volume of wet sediment recovered (1427-4049 m 3 ) as well as discharged (2693-6951 m 3 ) during the experiments, provide important inputs to evaluate the sediment resettlement and migration. Using these estimates, the average depth of excavation on the seafloor, can also been calculated. However, the stages of sediment recovery and discharge are expressed differently in some of the experiments. In order to standardize these, different stages have been identified, and definitions of certain terms have been suggested, for use in the future. The methods of calculating different properties, as well as weight and volume of discharged sediment are described in the paper for use in other applications concerning deep-sea discharges.     

胶州湾海水-沉积物界面氮、磷、铁的地球化学特征

对胶州湾底层水溶解氧、总氮、总磷、溶解无机氮、活性磷酸盐、铁及孔隙水中溶解无机氮、磷酸盐、铁和沉积物粒度、有机碳进行了分析。结果显示除铵氮外,孔隙水浓度明显高于底层水中浓度,其中硝酸氮、亚硝酸氮、活性磷酸盐、铁在孔隙水中的浓度分别为在底层水中浓度的17.8、9.3、12.5、7.7倍,暗示孔隙水中的物质可能向上覆水体扩散。在横向上,底层水及孔隙水中硝酸氮、亚硝酸氮、铵氮、活性磷酸盐都呈东岸高西岸低的分布规律,在西南部出现低值。Fe在底层水及孔隙水中的分布规律为东低西高,然而在沉积物中则与此正好相反。氮、磷、铁主要补给源有河流输入、工业生活污染排放、海洋生物自身分解以及孔隙水的释放。影响氮、磷、铁分布的主要因素为物源、河流输入及水动力条件,同时受到沉积物粒度的制约。相关分析显示,溶解氧、有机碳、铁对水体中磷及氮的分布具有某种制约作用。     

广东省的湿地:功能、价值、利用和湿地丧失的缓解措施

广东的湿地生态系统包括基塘系统、红树林、湿生植被和水生植被等4种生态系统，是特别而有经济价值的植被类型，这些湿地类型分别在支持生物地化循环、保堤护田护村、调节气候、净化环境、生物多样性保育、提供能源、提供生物学科学教育基地和生态旅游地、生产多样化的社会经济原料和产品等方面有着独特而重要的作用。但随着社会的发展．其面积在减少，质量不断下降。结构破坏直接导致了功能的下降和丧失。自1950年起，广东省在其湿地的保护和管理方面陆续实施了统一规划管理、设立自然保护区、开发湿地生产模式、防治湿地污染、保护湿地环境、建设人工湿地、恢复湿地系统、建立湿地信息系统等一系列举措，并取得了一定的收效，但效果仍不理想。要从根本上解决广东湿地的保护和利用问题，一方面要在恢复生态学的指导下深化退化湿地的恢复研究，按照湿地生态系统在空间尺度上的条带分布规律和时间尺度上的演替规律。因地制宜、因时制宜地进行湿地恢复研究；另一方面要进一步加强科研工作者、政府以及民众三者在湿地保护和合理利用方面的合作，建立专门机构，通过法律和行政手段进行统一规划、管理，同时对湿地保育研究、教育给予政策上的倾斜、支持。     

Carbon,nitrogen and phosphorus dynamics in nine sub-systems of the Sylt-Rømø Bight ecosystem,German Wadden Sea

Flow networks of nine sub-systems consisting of 59 components each of the Sylt-Rømø Bight, German Wadden Sea, were constructed depicting the standing stocks and flows of material and energy within and between the sub-systems. Carbon, nitrogen and phosphorous were used as currencies for each sub-system, thus resulting in 27 network models, which were analyzed by ecological network analytical protocols. Results show substantial variability in the dynamics of these elements within and between the nine sub-systems, which differ in habitat structure, species diversity and in the standing stocks of their constituent living and non-living components. The relationship between the biodiversity and selected information indices and ratios, derived from ecological network analysis, of individual sub-systems is variable and differ substantially between them. Ecosystem properties such as the structure and magnitude of the recycling of these elements, number of cycles, and total sub-system activity were calculated and discussed, highlighting the differences between and complexity of the flow of C, N and P in a coastal marine ecosystem. The average number of cycles increase from 179 for C, to 16,923 and 20,580 for N and P respectively, while the average amount of recycled material, as measured by the Finn Cycling Index (FCI), increase from 17% for C, to 52% for P and to 61% for N. The number of cycles and the FCI vary considerably between the sub-systems for the different elements. The largest number of cycles of all three elements was observed in the muddy sand flat sub-system, but the highest FCIs were computed for both C (32%) and N (85%) in the Arenicola Flats, and in sparse Zostera noltii sea grass beds for P (67%). Indices reflecting on the growth, organization and resilience of the sub-systems also showed considerable variability between and within the inter-tidal ecosystems in the Bight. Indices such as, for example, the relative ascendency ratios increase on average from C to N to P, whereas others, such as the Average Mutual Information and Flow Diversity indices, were found to be higher in the N models than in the C or P ones.     

Sediments of lakes Rotoroa and Rotoiti,South Island,New Zealand

The chemistry and mineralogy of the sediments of Lakes Rotoroa and Rotoiti, South Island, New Zealand (41° 51 S, 172° 38 E and 41° 50 S, 172° 50 E respectively) , were studied. In general, the cation exchange properties, element concentrations, and mineral compositions of the sediments show little variation between samples from each lake. However, some nutrient concentrations, notably exchangeable calcium, magnesium, and potassium, and total nitrogen and sulphur, are higher in sediments from weed beds than in sediments underlying deeper water; iron and manganese concentrations tend to show the opposite trend. Concentrations of exchangeable calcium and total nitrogen, sulphur, manganese, and iron generally decrease down the sediment profiles from the sediment/water interface. Differences between lakes, notably higher concentrations of illite and of the elements potassium, aluminium, titanium, and zirconium, and lower concentrations of calcium in the sediments of Lake Rotoiti than in those from Lake Rotoroa, are probably due mainly to the different geological settings rather than to differences in the diagenetic processes operating within the lakes. Mineralogical analyses indicate that the lake sediments are derived mainly from the valleys of the major inflow rivers rather than from the slopes at the sides of each lake.