Otolith microchemical evidence revealing multiple spawning site origination of the anadromous tapertail anchovy (Coilia nasus) in the Changjiang (Yangtze) River Estuary

The estuarine tapertail anchovy(Coilia nasus) is a high-value commercial fish. Estimating the spawning site or hatchery origin and habitat is essential for its conservation. This study aimed to determine the habitat use and life history characteristics of C. nasus from the Changjiang River Estuary. We investigated the environmental signatures of strontium(Sr) and calcium(Ca) in the otoliths of the collected specimens using electron probe microanalysis; additionally, we examined their gonadal mat...     

刀鲚、凤鲚和湖鲚矢耳石的形态学比较研究

选取凤鲚(Coilia mystus)、湖鲚(Coilia nasus taihuensis)以及采自崇明、黄海和瓯江口3个水域的刀鲚(Coilia hasps)的矢耳石为对象,描述了这些鱼类耳石的形态特征,并进一步使用基于12组形态数据的框架测量进行了种间和种内比较和判别分析.结果表明耳石形态的差异种间较种内更为明显...     

基于EPMA的耳石Sr：Ca比分析及其在鱼类生活履历反演中的应用实例研究

利用基于电子探针（EPMA）的耳石Sr：Ca比和Sr含量分析方法研究了长江口水域刀鲚、凤鲚、带鱼和长吻鲍的生活履历及生活史型。结果发现,刀鲚中除存在淡海水洄游性个体生活史型外。还存在出生并生活于河口或近海的非洄游性个体生活史型,其平均耳石Sr：Ca比在不同水环境履历的基准值为：淡水〈2．0×10^-3、河口（3．5—6...     

Comparing different spatial interpolation methods to predict the distribution of fishes: A case study of Coilia nasus in the Changjiang River Estuary

Spatial-temporal distribution of marine fishes is strongly influenced by environmental factors. To obtain a more continuous distribution of these variables usually measured by stationary sampling designs, spatial interpolation methods(SIMs) is usually used. However, different SIMs may obtain varied estimation values with significant differences, thus affecting the prediction of fish spatial distribution. In this study, different SIMs were used to obtain continuous environmental variables(water depth, water temperature, salinity, dissolved oxygen(DO), p H,chlorophyll a and chemical oxygen demand(COD)) in the Changjiang River Estuary(CRE), including inverse distance weighted(IDW) interpolation, ordinary Kriging(OK)(semivariogram model: exponential(OKE),Gaussian(OKG) and spherical(OKS)) and radial basis function(RBF)(regularized spline function(RS) and tension spline function(TS)). The accuracy and effect of SIMs were cross-validated, and two-stage generalized additive model(GAM) was used to predict the distribution of Coilia nasus from 2012 to 2014 in CRE. DO and COD were removed before model prediction due to their autocorrelation coefficient based on variance inflation factors analysis. Results showed that the estimated values of environmental variables obtained by the different SIMs differed(i.e., mean values, range etc.). Cross-validation revealed that the most suitable SIMs of water depth and chlorophyll a was IDW, water temperature and salinity was RS, and p H was OKG. Further, different interpolation results affected the predicted spatial distribution of Coilia nasus in the CRE. The mean values of the predicted abundance were similar, but the differences between and among the maximum value were large. Studies showed that different SIMs can affect estimated values of the environmental variables in the CRE(especially salinity).These variations further suggest that the most applicable SIMs to each variable will also differ. Thus, it is necessary to take these potential impacts into consideration when studying the relationship between the spatial distribution of fishes and environmental changes in the CRE.     

利用微阵列基因芯片研究东海海参(Acaudina leucoprocta)的生理功能

取喂食东海海参60d的ICR小鼠肝脏,制备微阵列基因表达芯片,并与模式小鼠芯片进行杂交,筛选功能基因和差异表达蛋白.从食用海参的鼠肝中共筛选出了4180个基因,其中差异表达基因184个,上调基因107个,下调基因77个.利用MAS.2.0系统对其功能进行分析,结果显示,分子功能上有催化活性的基因占39.86%,结合功能...     

泥蚶低氧胁迫后血细胞转录组及耐低氧相关基因的筛选与分析

泥蚶(Tegillarcagranosa)是一种比较特别的具有较强耐低氧能力的贝类,但目前其耐低氧分子调控机理尚不知。为探究泥蚶耐受低氧的分子调控机理,本研究对低氧胁迫下的泥蚶(Tegillarca granosa)血细胞转录组中富集的信号通路及生物学过程进行分析,并初步筛选分析耐低氧基因。对低氧(DO=0.5 mg/L)胁迫6、24、72、120 h的泥蚶血细胞进行转录组测序(RNA-seq)并开展生物信息学分析,筛选出差异基因集,并对胁迫72、120 h的DEGs进行GO富集分析、KEGG通路分析,采用qPCR方法检测7个DEGs的表达量并与转录组比较。结果显示从6 h开始到120 h的4个时间点的DEGs数量呈增多的趋势, GO功能分析主要富集在JUN激酶活性的负调控、蛋白质水解的负调控及免疫系统进程等主动抗凋亡、抗逆进程; KEGG通路分析主要富集在胰岛素及胰腺分泌的信号通路、HIF-1通路、钙信号相关通路和细胞凋亡相关通路。qPCR检测结果显示, 7个基因的表达上调/下调趋势与转录组测序一致,证实了转录组测序结果的可靠性。本研究推测胰腺分泌信号通路、钙信号通路及凋亡通路在泥蚶...     

高温胁迫下坛紫菜的数字基因表达谱研究

坛紫菜是潮间带重要的经济藻种,对高温、渗透压等逆境具有独特的调控机制。本文采用基于高通量测序的数字基因表达谱(DGE)技术研究了坛紫菜在高温胁迫下的基因表达差异,并分析其相应的响应方式;利用实时定量PCR技术对DGE部分数据进行验证;检测了其中较有代表性的应答基因hsp70的差异表达。结果显示,高温胁迫下坛紫菜中有256个unigene上调表达,以HSP、核糖体蛋白L12、延伸因子EF-Tu及部分光合作用相关基因为代表,3 820个unigene下调表达,主要为核酸、蛋白以及糖类等合成代谢相关基因。Gene Ontology分析表明,差异表达基因主要定位于质体等有膜细胞器,参与繁殖和发育过程,行使催化和连接酶活性的功能。Pathway分析显示,这些基因分布于107条pathway中。其中,下调表达基因最显著富集于mRNA监督和RNA转运途径,而上调表达基因部分富集于内质网的蛋白加工、RNA降解及光合作用途径。验证表明此次DGE结果具有较高准确性,hsp70基因对高温响应积极。综上所述,DGE结果反应出,在高温胁迫时,坛紫菜出现基础代谢减慢、合成速度下降、能量合成受阻、碳同化降低等现象,但光合作用前期未受影响,同时补救途径启动。     

α2,8-唾液酸转移酶Ⅵ过量表达影响小鼠乳腺癌细胞基因表达的研究

研究α2,8-唾液酸转移酶VI(Sixth type of α2,8-sialyltransferase, ST8Sia VI) 对乳腺癌细胞生物学功能的影响及其作用的分子机制.采用全基因组芯片技术检测ST8Sia VI过表达前后小鼠乳腺癌细胞4T1基因表达谱的差异;利用PathwayExplorer分析ST8Sia VI过表达前后对基因网络的影响.实验结果表明:ST8Sia VI过表达引起201个基因表达有差异,其中22个基因与肿瘤细胞的黏附、生长、运动、免疫和周期有关.另外,PathwayExplorer分析结果显示,差异表达基因涉及的最显著变化的基因网络是"依赖β-arrestin 的G蛋白偶联受体(GPCR)信号通路";进一步的实验结果证明该通路下游Raf蛋白的磷酸化水平在ST8Sia VI过表达细胞显著升高.上述结果为ST8Sia VI生物学功能的深入研究提供了前提.     

Identification of transcriptional effects of ethynyl oestradiol in male plaice (Pleuronectes platessa) by suppression subtractive hybridisation and a nylon macroarray

Suppression subtractive hybridisation (SSH) was used to generate cDNA libraries representing genes differentially expressed in response to ethynyl oestradiol (EE2) exposure in liver from male plaice (Pleuronectes platessa) previously analysed for vitellogenin (VTG) induction. Characterisation of the cDNA clones identified many as VTG (2 genes) and zona radiata proteins (ZRP) (3 genes), but 40 encoded other proteins, with more than half cryptic. Further analysis identified 85 non-redundant clones suitable for array on nylon membrane. Radiolabelled cDNAs were prepared from hepatic mRNA from EE2 treated plaice (0 and 21 days) and hybridised with the arrayed clones. Analysis of the data showed that 11/17 novel, 21/22 VTG, 13/14 ZRP, 2/2 liver aspartic proteinase (LAP) and 8/10 other mRNAs were up-regulated by EE2 exposure.     

泥蚶(Tegillarca granosa)肉质的蛋白组学研究

采用双向电泳技术,对广东阳江和浙江乐清两地的泥蚶肌肉全蛋白进行了研究,并利用质谱和软件对差异蛋白进行了分析.研究结果表明,阳江和乐清两地样品平均检测到的蛋白点数分别为674±25和746±34个,平均匹配率为78.6%.差异表达蛋白质点数为26个,其中3个点仅在阳江泥蚶中表达,5个点仅在乐清泥蚶中表达;6个点在阳江泥蚶中为高表达,12个点在阳江泥蚶中为低表达.选择其中6个具有代表性的蛋白点进行肽指纹图谱分析.结果显示,乐清泥蚶中高表达的4个蛋白确定为肌动蛋白、PRKA激酶锚定蛋白9、烯醇化酶.这些物质与泥蚶的肉质特性有一定关联.乐清地区出产的泥蚶在营养、肉质和鲜美度方面均优于阳江地区的泥蚶.     

Identification of transcriptional effects of ethynyl oestradiol in male plaice (Pleuronectes platessa) by suppression subtractive hybridisation and a nylon macroarray

Suppression subtractive hybridisation (SSH) was used to generate cDNA libraries representing genes differentially expressed in response to ethynyl oestradiol (EE2) exposure in liver from male plaice (Pleuronectes platessa) previously analysed for vitellogenin (VTG) induction. Characterisation of the cDNA clones identified many as VTG (2 genes) and zona radiata proteins (ZRP) (3 genes), but 40 encoded other proteins, with more than half cryptic. Further analysis identified 85 non-redundant clones suitable for array on nylon membrane. Radiolabelled cDNAs were prepared from hepatic mRNA from EE2 treated plaice (0 and 21 days) and hybridised with the arrayed clones. Analysis of the data showed that 11/17 novel, 21/22 VTG, 13/14 ZRP, 2/2 liver aspartic proteinase (LAP) and 8/10 other mRNAs were up-regulated by EE2 exposure.     

青蛤(Cyclina sinensis)IKK基因的克隆及其在鳗弧菌(Vibrio anguillarum)刺激下的表达分析

利用构建的青蛤(Cyclina sinensis)转录组文库,筛选到青蛤IKK基因的类似序列。经设计引物克隆比对后确认为CsIKK基因。利用生物信息学软件在线对该基因进行结构分析。采用PCR技术克隆基因,并使用实时荧光定量PCR技术克隆得到CsIKK基因在青蛤五个不同组织中的表达情况及在鳗弧菌(Vibrioanguillarum)的刺激下IKK基因在青蛤血淋巴中的时序性表达情况。综合结果得到, CsIKK基因序列开放阅读框长2298bp,编码765个氨基酸。IKK基因在青蛤的血淋巴、外套膜、闭壳肌、肝脏、性腺和鳃六个组织中均表达,在血淋巴中表达量最高。青蛤IKK基因在鳗弧菌胁迫下表达量在6h时达到最大值,与对照组相比差异极显著(P0.01),表明该基因所指导的蛋白是青蛤重要的免疫信号通路蛋白。     

松江鲈肌酸激酶的基因克隆、表达与功能分析

前期蛋白质质谱分析发现,松江鲈（Trachidermus fasciatus Heckel）经鳗弧菌（Vibrio anguillarum）刺激后,其皮肤黏液中的肌酸激酶（creatine kinase,CK）的表达明显上调。目前,肌酸激酶在鱼类中的功能研究尚未深入开展。本文利用RACE技术克隆获得了松江鲈CK基因的全长cDNA序列（命名为TfM-CK）。TfM-CK基因的cDNA全长为1 474 bp,开放阅读框为1 146 bp,编码381个氨基酸。序列比对分析显示,TfM-CK序列高度保守。实时荧光定量PCR显示,TfM-CK mRNA广泛表达于松江鲈各组织,在肌肉中的相对表达量最高,其次为血液。鳗弧菌刺激后,TfM-CK mRNA在肌肉、皮肤、脾脏和头肾中均上调表达。其中,脾脏中的表达量高达对照组的900多倍。原核重组表达纯化的TfM-CK （rTfM-CK）蛋白酶活测定实验结果显示,重组蛋白的酶活为22.0 U/mg。rTfM-CK对鳗弧菌、大肠杆菌（Escherichia coli）、巨大芽孢杆菌（Bacillus megaterium）和金黄色葡萄球菌（Staphylococcus aureus）有较强的凝集作用。由以上实验结果推测,TfM-CK可能通过细菌凝集作用参与到了松江鲈抵抗病原菌的先天免疫防御过程中。本研究为认识鱼类肌酸激酶的功能及其在病原菌防御过程中的分子免疫机制机制提供了参考。     

Differential gene expression in Poecilia vivipara exposed to diesel oil water accommodated fraction

Diesel fuel is a potential contaminant of estuarine and mangrove areas, particularly because it is the main fuel used in small boats and larger vessels. The aim of this work was to identify genes differentially expressed in the liver of Poecilia vivipara (Guppy) exposed to 10% diesel fuel water accommodated fraction (WAF), employing the subtractive suppressive hybridization (SSH) method. The results showed 27 differentially expressed gene fragments, 12 up-regulated and 15 down-regulated. Among the up-regulated genes were CYP1A, UDPGT1a, ABCC4, Methyltransferase and Apolipoprotein A1. Down-regulated genes included Vitellogenins, C1 Inhibitor and Complement Component 3c. The identified genes are associated with different metabolic functions like biotransformation, membrane transport and immune system, indicating the susceptibility and/or molecular responses of this organism to the toxic effects elicited by diesel fuel WSF.     

转录组学分析B-3胞外多糖对巨噬细胞RAW264.7基因表达的影响

B-3胞外多糖是从一株南极嗜冷菌Psychrobacter sp.B-3发酵液中提取的胞外多糖,前期研究发现该多糖能够激活巨噬细胞并影响其免疫调控活动。为探明其免疫调控途径,采用转录组学分析对经B-3多糖刺激的RAW264.7巨噬细胞系进行了差异表达基因分析。研究结果表明B-3多糖的刺激引起了420个基因的差异表达,其中上调178个、下调242个。差异表达基因与细胞功能存在诸多关联,但主要集中在代谢和免疫两个通路上：其中在多个免疫相关通路上,富集最显著的是抗原处理和表达途径;另一个明显富集的则是代谢通路,B-3多糖处理引起了大量参与代谢的酶类的表达变化,包括氨基酸,碳水化合物,脂质和核苷酸代谢相关基因均发生了一定程度的改变。因此,B-3胞外多糖作为一种免疫激活剂直接影响了RAW264.7巨噬细胞系的免疫功能,同时也对其胞内的代谢途径产生了影响。本论文首次开展了南极嗜冷细菌胞外多糖对RAW264.7巨噬细胞系的影响,该研究为不同来源的活性多糖对巨噬细胞免疫激活研究提供了科学依据。     

基于马氏珠母贝(Pinctada fucata)血细胞全长转录组数据补体样组分的鉴定与分析

补体系统作为先天免疫的重要组成部分,是一种复杂的限制性蛋白水解系统,其在免疫系统中发挥着重要的防御作用。为分析马氏珠母贝补体系统的组成及作用机制,使用血细胞样品进行了全长转录组测序建库、基因比对、功能注释,共挖掘到212个潜在补体样组分相关基因。补体样组分基因经同源性比对和结构域检测分析表明,检索到的基因分别编码89个含C1q结构域蛋白、57个C型凝集素蛋白、33个纤维胶凝蛋白、11个纤维蛋白原相关蛋白、8个甘露糖结合型凝集素关联丝氨酸蛋白酶、2个含硫酯蛋白(1个C3分子,1个TEP分子)、1个补体受体、2个补体因子、9个丝氨酸蛋白酶。随机选择12个补体相关基因,使用溶藻弧菌刺激前后的血细胞样品进行实时定量PCR检测其表达水平,结果显示C1q(C1q domain containing protein)、C-lectin、MBL(mannose-binding lectin)、ficolin、MASP(mannan-binding lectin serine protease)等基因均呈现出显著差异表达,表明马氏珠母贝补体系统是一个复杂的多组分效应系统,且可能通过凝集素途径或类似于凝集素途径激活补体系统的免疫作用。研究结果为进一步验证马氏珠母贝中存在的原始补体系统提供了分子生物学证据,同时对深入了解马氏珠母贝免疫防御机制,丰富和发展海洋无脊椎动物免疫学内容也具有重要理论意义。     

Reproductive periodicity in Evechinus chloroticus in the Hauraki Gulf

Test diameter and gonad volume were recorded over 1 year for bimonthly samples of Evechinus chloroticus (Valenciennes) from each of 3 subtidal populations 10 km apart in the Hauraki Gulf. Histological analysis was used to relate cellular events during gametogenesis to the annual cycle of change in gonad size. Mean gonad sizes of the different sea urchin populations were compared by analysis of covariance. Gametogenesis began in the spring, and major spawning occurred in mid to late summer. The proportion of the gonads taken up by nutritive phagocyte cells increased from autumn to spring. During proliferation and growth of gametes the nutritive phagocytes declined in abundance and globulation, suggesting that reserves stored in these cells were transferred to developing gametes. Gonad size doubled during gametogenesis, declined after spawning, and remained low in autumn and winter. Differences in gonad size between populations were significant during most of the year, and were most pronounced in mid summer. Although synchronous gametogenesis occurred in the 3 populations studied, spawning occurred at different times. This suggested that spawning in this species was induced by factors acting either within sea urchin populations or over distances of a few km or less.