Zoospore-derived monoecious gametophytes in Undaria pinnatifida (Phaeophyceae)

The annual life cycle of the brown seaweed Undaria pinnatifi da(Harvey)Suringer comprises a macroscopic diploid sporophyte stage and a microscopic haploid gametophyte stage.In 2011,an unusual zoospore-derived monoecious gametophyte isolate(designated as line 10-5-3)of U.pinnatifi da was observed.To understand this phenomenon,a comprehensive screening of eighty-two previously identifi ed male gametophyte cultures,isolated from three randomly selected cultivars(lines 10,7,and 5)was performed.Thirty-six of the isolates developed both antheridia and oogonia on the same fi lamentous fragment in a standard gametogenesis test(SGT: 18°C,60 μmol photons/(m 2·s)).Selfi ng of the monoecious gametophyte or crossing it with a normal male gametophyte both gave rise to morphologically normal sporophytic offspring.However,crossing resulted in a much higher fertilization rate(89.7%).The hybrid and selfed sporophytic offspring were grown to maturity in fl ow tanks at an ambient temperature of 10–18°C over a period of 69 days.Active zoospores were released from both types of mature sporophylls.The majority of these developed into male gametophytes,while 15%–20% developed into the observed monoecious structures on the same fi lament.Using PCR amplifi cation it was found that all the monoecious gametophyte isolates and the sporophytic offspring resulting from the selfi ng and crossing lacked the femalelinked microsatellite sequence(a part of the locus Up-AC-2A8,GenBank accession No.AY738602.1),indicating their male nature.U.pinnatifi da is an invasive species in some regions and the implications of the above fi ndings for this species in nature are briefl y discussed.     

溶藻弧菌acfA基因克隆与生物信息学分析

溶藻弧菌引起海水养殖产业弧菌病的主要病原,附着定植因子基因acfA是溶藻弧菌重要毒力基因之一,其表达产物是溶藻弧菌有效定植在宿主肠道上所必需的蛋白。根据弧菌属细菌acfA基因基因保守区设计引物,采用Touchdown PCR扩增acfA基因部分序列,Inverse PCR和Nested PCR扩增已知序列侧翼序列,成功克隆了溶藻弧菌acfA基因全长。克隆的acfA基因序列全长为743 bp,开放阅读框长度为648 bp组成,共编码215个氨基酸,在5′端上游未发现-35区和-10区序列。演绎的ACFA蛋白N端有18个氨基酸信号肽序列,表明该蛋白是分泌型蛋白;蛋白结构分析表明主要由α螺旋和不规则卷曲构成。BLAST分析表明,该蛋白氨基酸序列与其他弧菌相应蛋白同源性较高,是较保守的外膜蛋白。     

三疣梭子蟹(Portunus trituberculatus)胞内氯离子通道蛋白基因克隆及其表达分析

本研究克隆了三疣梭子蟹(Portunus trituberculatus)胞内氯离子通道蛋白基因,命名为Pt CLIC。该基因c DNA全长2000bp,5’和3’非编码区(UTR)长分别为76bp和1162bp,开放阅读框长762bp,推测编码253个氨基酸,预测分子量为29.2k Da,理论等电点为5.93。生物信息学分析表明,Pt CLIC基因中未发现跨膜结构域,属于不稳定蛋白;同源性分析表明,Pt CLIC基因编码的氨基酸序列与蚤状溞(Daphnia pulex)CLIC基因的同源性高达83%;系统进化分析表明,三疣梭子蟹与拟穴青蟹首先聚为一支;实时荧光定量RT-PCR分析表明,Pt CLIC基因在选取的所有组织中均有表达,在肝胰腺中的相对表达量最高,且显著高于其它组织(P0.05)。低盐度胁迫显著改变了Pt CLIC基因在三疣梭子蟹鳃和肝胰腺中的表达模式,整体呈先上调后下调的趋势,并发现该基因在低盐耐受和低盐敏感家系中的表达规律差异显著。研究结果表明Pt CLIC基因在三疣梭子蟹渗透压调节中发挥重要作用,可辅助三疣梭子蟹耐低盐品系的选育。     

Cloning and expression analysis of the chloroplast fructose- 1,6-bisphosphatase gene from Pyropia haitanensis

Fructose-1,6-bisphosphatase(FBPase) is one of the key enzymes in Calvin circle and starch biosynthesis. In this study, the full-length of cpFBPase gene from Pyropia haitanensis was cloned by using rapid amplification of cDNA ends(RACE) technology. The nucleotide sequence of PhcpFBPase consists of 1 400 bp, including a 5′ untranslated region(UTR) of 92 bp, a 3′?UTR of 69 bp, and an open reading frame(ORF) of 1 236 bp, which can be translated into a 412-amino-acid putative peptides with a molecular weight of 44.3 kDa and a theoretical pI of 5.23. Multiple sequence alignment indicated that the protein belonged to the chloroplast FBPase enzyme. Phylogenetic analysis showed that the protein assembled with the cpFBPase of a thermal tolerant unicellular red micro-algae Galdieria sulphuraria. Expression patterns analyzed by qRT-PCR revealed that the expression of PhcpFBPase gene in the thallus phage was 7-fold higher than in the conchocelis phage, which suggested the different mechanisms of inorganic carbon utilization among the different life phages of P. haitanensis. And the different response modes of PhcpFBPase mRNA levels to high temperature and desiccation stress indicated that PhcpFBPase played an important role in responsing to abiotic stress.     

牙鲆NPY的体外表达及体内检测

神经肽Y(Neuropeptide Y,NPY)被普遍认为是一种重要的促食因子,在调节鱼类的摄食行为方面起着至关重要的作用。为进一步研究牙鲆NPY蛋白的生物学功能,作者克隆了牙鲆NPY成熟肽序列(m-NPY)及含有信号肽的全长序列(f-NPY),利用原核表达系统分别进行体外重组表达,筛选出诱导剂IPTG最佳诱导浓度为0.8 mmol/L及最佳诱导时间3 h;此外,根据牙鲆NPY第49-64位氨基酸序列制备了多克隆抗体并通过Western blot验证该多抗能够有效检验重组NPY及牙鲆体内NPY的表达。研究结果为研究重组牙鲆NPY蛋白在牙鲆水产养殖产业中的应用及检测提供了依据。     

基于遗传克隆选择算法优化BP神经网络的地理信息预测研究

针对BP神经网络训练过程中的训练时间较长、完全不能训练或容易陷入局部极小值等问题,提出基于遗传克隆选择算法(CLOGA)优化BP神经网络的流程,克服BP算法的一些缺陷。并通过湖北省人口预测问题进行效果检验,得到满意的结果。     

元胞自动机和克隆选择算法的蓝藻水华预测模型

针对蓝藻水华频发影响水体质量和生态环境的问题,该文以太湖为例,探讨了元胞自动机和克隆选择算法在蓝藻水华预测方面的可行性。使用遥感经验算法计算水体叶绿素a浓度;将叶绿素a浓度分布数据作为预测模型的输入数据,将基于平衡方程的预报模型与元胞自动机进行耦合,对蓝藻水华时空变化过程进行了动态模拟;在预测方面实现了蓝藻水华空间分布情况的可视化,并采用克隆选择算法优化了预测模型中的重要参数。研究结果表明:利用克隆选择算法优化的元胞自动机模型能够在蓝藻水华预测中直观地显示蓝藻水华的时空变化特征;能够在短时间内重复性地预演蓝藻水华的变化过程,为决策者提供有用的参考。     

一株H_5N_1亚型禽流感病毒株NA基因的克隆与序列分析

用RT-PCR方法从1个H5N1亚型禽流感病毒分离株A/Chicken/Guangdong/DH/1997扩增NA基因cDNA片段,将其克隆至pMD18-T载体,获得重组质粒pMD-NA,并对其核苷酸序列进行测定和分析。结果表明,该毒株的NA基因长度为1350bp,编码449个氨基酸,与其它H5N1亚型AIV分离株的核苷酸序列同源性为97.0%～99.4%,氨基酸序列同源性为97.7%～99.1%,提示禽流感病毒NA基因保守性较高。NA基因氨基酸序列的聚类分析表明该毒株与来自香港的A/Pheasant/HK/FY155/01和A/Ch/HK/FY150/01两个分离株处于同一进化枝,亲缘关系较近。     

裙带菜配子体对三种重金属离子毒性耐受力的研究

裙带菜于１９９４年采自威海市俚岛养殖场。１９９５年２月－１９９７年３月,用不同浓度（０,１,１０,１００,１０００μｍｏｌ／Ｌ）的Ｃｕ＾２＋,Ｃｄ＾２＋和Ｚｎ＾２＋处理裙带菜配子体,研究这些重金属对其生长和代谢的影响。结果表明,Ｃｕ＾２＋对裙带菜配子体生长的抑制效应最强：在１μｍｏｌ＋Ｌ时已出现抑制效应,叶绿素含量和光合作用速率明显下降,脯氨酸和丙二醛含量升高;１０μｍｏｌ／Ｌ时,配子体生长几乎停     

鲨肝刺激物质类似物基因在大肠杆菌中的表达与产物活性分析

构建了鲨肝刺激物质类似物基因片段的原核表达载体质粒 (pET-sHSS) ,转化大肠杆菌后通过IPTG诱导获得原核表达产物 ,并利用凝胶层析方法对表达产物进行了分离纯化 ;进一步利用MTT比色法研究了该重组产物对肝癌细胞株SMMC -7721的增殖影响。结果表明 ,在1mmol/L的IPTG的诱导下 ,鲨肝刺激物质类似物基因片段在大肠杆菌BL21菌株中获得表达 ,表达产物的相对分子质量大小约为17500u,占菌体可溶性蛋白总量的38%左右 ;纯化后的产物在低浓度下 (<50ng/L)具有明显刺激SMMC7721细胞株增殖的活性 ,高浓度下 (>100ng/L)则明显抑制该肿瘤细胞株的生长 ,与天然鲨肝刺激物质的生物活性类似