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~

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

Biochemical compositions of Tachypleus tridentatus at embryonic development stage

国家自然科学基金资助项目! ( 3 9670 147

Separation and Identification of β-alanine in Sargassum horneri(Turn.) C.Ag.

非蛋白质氨基酸在抗癌、抗菌、抗结核、抗坏血病等方面有着重要的作用。本文主要对铜藻中的游离氨基酸进行检测分析和部分分离纯化及结构鉴定方面的研究,为更好的开发利用这些天然产物提供技术支持。采用离子交换树脂层析法分离铜藻粗提液中的游离氨基酸,收集3 MOl/l的氨水洗脱液,用PITC-HPlC柱前衍生反相高效液相色谱法对其进行检测分析,结果显示粗提液中除含有多种组成蛋白质的氨基酸外还有3种未知组分,且含量较高的常见蛋白质氨基酸为丙氨酸、脯氨酸、缬氨酸。采用半制备高效液相色谱系统制备分离了其中一种未知组分的衍生物MWz2,经真空冷冻干燥后为略黄固体粉末,结合核磁共振波谱、高分辨质谱、红外光谱数据,最终鉴定MWz2去掉已知取代基团PITC后的成分为β-丙氨酸,分子式为C3H7nO2,分子量为89.09。Non-protein amino acids were found to have many bioactivities such as anti-cancer,antibacterial,anti-tuberculosis,anti-scurvy and so on.Detection,separation and identification of free amino acids in Sargassum horneri(Turn.) C.Ag.were studied in this paper,which provided technical support for development and utilization of these natural products.Free amino acids were separated by ion-exchange column chromatography from the crude extracts of Sargassum horneri(Turn.) C.Ag.Amino acids were eluted with 3 mol/L NH4OH and the eluate was collected.Many protein amino acids and three unknown components were discovered in Sargassum horneri(Turn.) C.Ag.by PITC-HPLC.The main protein amino acids were alanine,proline,valine.One unknown derivative component(MWZ2) was obtained by preparation HPLC.MWZ2 was slightly yellow solid powder after vacuum freeze-drying.According to analysis of NMR,HR-MS,IR spectroscopy data,the molecular structure of MWZ2 had been elucidated.MWZ2 was β-alanine after removed substituent component of PITC,its molecular formula was C3H7NO2,molecular weight was 89.09.国家海洋局海洋公益项目(200805041

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

绿色荧光蛋白基因在大黄鱼体内的表达

以绿色荧光蛋白基因为报告基因 ,以pIRESneo为载体质粒 ,制备含有报告基因的质粒 ,以肌肉注射的方式导入大黄鱼体内 ,每隔一周用荧光显微镜观察绿色荧光蛋白基因在鱼体不同部位的表达情况 .结果表明 ,绿色荧光蛋白基因可以在大黄鱼体内得到表达 ,表达时间高达 2 8d以上 .绿色荧光蛋白基因不仅可以在肌肉注射部位组织细胞得到表达 ,而且在其它部位的肌肉、肝脏、肾脏和心脏等组织中也有表达 ,但表达水平低于肌肉注射部位组