利用改良Nest-tetra-primerspecificPCR技术对两种石斑鱼的鉴别

云纹石斑鱼(EPInEPHEluS MOArA)和褐石斑鱼(E.brunEuS)为石斑鱼属内近缘种,外形相似常被混淆.本研究改良建立了nEST-TETrA-PrIMEr SPECIfIC PCr方法,获得了云纹石斑鱼和褐石斑鱼线粒体dnAnd2基因内的3个特异性条带,分别为内参序列nC1(394bP)、特异性条带nd2-M(268bP)和nd2-b(122bP),以及核基因组中核糖体dnAITS1区的5个特异性条带,分别为内参序列nC2(588bP)、nC3(563bP),特异性条带rdnA-M(426bP)、ITS1-M(488bP)和ITS1-b(304bP).研究结果不仅为两种石斑鱼的鉴别提供了稳定、可靠、快捷的特异性分子标记,而且也为鱼类近缘种的dnA鉴别提供了新的途径.国家自然科学基金(批准号:40576064);国家“八六三”计划(项目编号:2006AA10A414)资助项

三种海洋浮游甲壳动物Tm的研究

纯化三次的日本毛虾、中国毛虾和中华假磷虾的Tm含量分别占其干重的0.86％,0.87％,和0.70％;SDS-PAGE测定两种毛虾Tm亚基分子量约3.3—3.6×104,磷虾的约3.2—3.5×104;等电聚焦分析出此三种动物Tm的pI约为pH5.0,在pH4.8或7.2的镁溶液中,三种Tm均能形成针状晶体,透射电镜下可见到约39nm横纹周期及若干细节;三种Tm均含有Glu,Asp、Lys、His,Arg、Ser、Thr、Gly、Ala、（Cys）2、Val、Met、Ile、Leu、Tyr和Phe,但都未测出Pro和Trp;圆二色性测定指出,在远紫外区三种Tm均显示出峰型极为相似的典型α螺旋双负峰,峰极点分别落在222和208nm处,日本毛虾、中国毛虾和中华假磷虾Tm在208nm处的α螺旋度分别占整个肽链的76.6％,74.2％和61.6％

基于枯草芽孢杆菌重组菌株构建技术制备海洋源金属硫蛋白

为构建安全高效表达海洋源金属硫蛋白的益生菌重组菌株,以海洋源褶牡蛎内脏团为模板,PCR克隆得到褶牡蛎金属硫蛋白基因(Op-MT),大小为324 bp,经pMD19-T载体亚克隆到融合表达载体pHT43-SUMO中,然后通过电转化法将重组质粒转入八种蛋白酶缺陷的宿主枯草芽孢杆菌(Bacillus subtilis)WB800N中。SDS-PAGE电泳显示诱导表达的融合蛋白SUMO-Op-MT分子量约为23 kD,与预期一致,说明可溶性融合蛋白SUMO-Op-MT在宿主菌中得到了表达,且通过Western blotting进一步得到验证。同时,HPLC检测重组枯草芽孢杆菌发酵液,结果也表明重组枯草芽孢杆菌成功表达了可溶性目的蛋白Op-MT。通过对重组工程菌中MT活性检测发现,其对重金属镉耐受力增强,说明重组工程菌成功表达出有活性的金属硫蛋白。这为以后开发天然、高效海洋源重金属解毒剂提供理论基础。福建省科技计划项目(2017N0016);;国家海洋局第三海洋研究所基本科研业务费专项资金资助项目(海三科2016031)共同资

Bioinformatics analysis of QM-like(PcQM) protein from Pseudosciaena crocea

[目的]通过分析PCQM蛋白质的结构,预测其高级结构和功能,为进一步研究PCQM蛋白提供理论依据。[方法]利用blAST P和相关分析蛋白质理化性质与结构的软件,对PCQM蛋白进行结构与功能预测分析。[结果]PCQM蛋白不存在n端信号肽,存在多个酶切位点,无跨膜结构,定位在细胞质中。不存在二硫键,该蛋白含26.05%α-螺旋结构(H),20.93%β-折叠结构(E),53.02%无规则卷曲(l)。最后,通过SWISS-MOdEl程序预测了PCQM蛋白的三级结构。[结论]较全面地分析和预测了PCQM蛋白的理化性质、二级结构和三级结构等。为深入分析PCQM在大黄鱼(lArIMICHTHyS CrOCEA)体内免疫调节功能研究,提供了理论依据。[Objective]The purpose of this study was to learn more function of PcQM protein by analyzing the AA sequence and predicting the structure of it.[Methods]Some related bioinformatics analysis software or system on internet such as Blast P,Predict Protein,Swiss-Model,was used to analyze the structure and prediction the structure and function of PcQM protein.[Results]The results showed that PcQM protein had many restriction enzyme sites.There was no signal peptide on N-terminal and no transmembrane structure.It was located in the cytoplasm.There were 26.05% α-helical(H),20.93% β-pleated sheet(E) and 53.02% random coil(L) predicted by Predict Protein.The tertiary structure was predicted by Swiss-Model.[Conclusion]The study has analyzed the sequence of PcQM protein and predicted the structure of it,which would lay a foundation for further learning the relationship between structure and function of the protein.江苏省高校自然科学研究面上项目(No.12KJD240001)资助~

The Karyotypes of Nibea diacanthus

以头肾细胞为材料,采用PHA体内注射,空气干燥法制片,分析了双棘黄姑(Nibeadiacanthus)的染色体组型.结果表明:其染色体组型为:2n=48=24t,NF=48.在已进行过染色体组型分析的石首鱼类中,除大黄鱼外,染色体组型均为2n=48t.The karyotypes of Nibea diacanthus were examined in the paper.The samples were separately injected by PHA and colchicines.The chromosomes were received from the renal tissue by using air drying method and Giemsa staining.The result showed that the chromosome number of Nibea diacanthus was 48;and its karyotype formula was 2n=48=48 t,NF=48;it was the same as the other croakers determined,except Pseudoceana crocea in Sciaenidae.福建省自然科学基金(B0110004)资助细胞数目938770

Study on protease activities of the ECP of pathogenic bacterium Vibrio harveyi strain TS-628

采用复性电泳技术,研究环境pH、温度及培养时间对哈维氏弧菌TS-628菌株胞外产物(ECP)蛋白酶活性的影响。结果表明,胞外产物蛋白酶最适反应pH在8.5左右,酸对ECP蛋白酶活性的抑制明显强于碱对它的抑制作用;最适反应温度在20～50℃之间,20℃以下或50℃以上蛋白酶活性则受到明显抑制,表明蛋白酶不仅对高温敏感,对低温也很敏感;培养12h的胞外产物蛋白酶活性最强。最适温度和pH条件下,主要出现3种蛋白酶,其中分子量为94kDa和26kDa的两种蛋白显示了很强的蛋白酶活性,而且能在比较极端的温度和pH值条件下保持活性,而分子量为35kDa蛋白酶只在最适反应条件下出现,并且该蛋白酶的最适反应条件与弧菌病暴发时的环境条件极为相似。由此可推测分子量94kDa和分子量为26kDa的两种蛋白可能是弧菌维持生存所必需,而分子量为35kDa的蛋白酶虽然只出现在特定条件下,却可能在致病过程中起重要作用。SDS-gelatin-PAGE assay was used to study the effects of pH,temperatures,and growing time on the protease activities of the extracellular products(ECP) of Vibrio harveyi TS-628 strain.The results showed that the optimal pH for ECP protease activity was about 8.5 and the protease activity was inhibited more easily by lower values of pH than by higher ones.The optimal temperature for the protease was 20~50℃,and significant decrease in activity was observed at temperatures below 20℃ or above 50℃,which indicated that the protease was sensitive to both low and high temperatures.The highest level of the extracellular product protease activity was present after 12h incubation.Under the condition of optimal temperatures and pH,there were three kinds of protease present:94kDa,26kDa and 35kDa.The 94kDa and 26kDa proteases displayed high activity and could remain active under extreme temperatures and pH,while the 35kDa protease was only active under the optimal condition,which was similar to the condition for the outburst of pathogenic vibrio.It is suggested that the 94kDa and 26kDa proteases were necessary in maintaining the vibrio survival and the 35kDa protease could play an important role in the pathogenicity of the bacteria.福建省青年科技人才创新基金项目(2006F3096);; 集美大学科研基金项目共同资

Embryonic Development of Tachypleus tridentatus

用人工授精方法获取中国鲎的受精卵 ,室温下完成胚胎发育。实验结果表明 ,①中国鲎成熟卵为黄色或浅黄色 ,卵径 2 7～ 3 5mm ,扫描电镜下厚而韧的卵壳上有许多大小不同的微孔 ,精子头部椭圆形 ,具一盘状顶体囊 ,尾呈鞭毛状 ;②中国鲎的胚胎发育可分为卵裂期、囊胚期、原肠胚形成期、附肢原基形成期、组织分化期和快速生长期 ,从受精到孵出三叶幼体约需 55d ;③中国鲎胚胎发育具有发育期长、个体差异大、部分胚胎发生滞育等特点。Mature horseshoe crab, Tachypleus tridentatus were sampled from aquatic market in Xiamen during the period from July 1998 to July 1999. The fertilized eggs were obtained by artificial insemination, then cultured in prepared seawater at room temperature(24～30℃). The fertilized egg appeared yellow or light yellow and its size ranged from 2.7 to 3.5*!mm. Sperm of the horseshoe crab consisted of an elliptical head and a long flagellum. Under the scanning electron microscope, there were many irregular pores on the thick chorion surface and an acrosomal vesicle like a plate on the top of the sperm. The embryonic development of T.tridentatus could be divided into 6 stages: cleavage,blastoderm,gastrule,appearance of rudimental appendages,organ specialization and rapid growing stage. It took about 55 days for the embryonic development from fertilized eggs to trilobite larva. The embryonic development of the horseshoe crab showed the characteristics of long hatching period,great variations in development interval between the individuals and diapause in some embryos.国家自然科学基金资助项目 (No.396 70 147

Biochemical compositions of Tachypleus tridentatus at embryonic development stage

国家自然科学基金资助项目! ( 3 9670 147