Construction of bacterial artificial chromosome libraries for Zhikong Scallop <Emphasis Type="Italic">Chlamys farreri</Emphasis>

Two Large-insert genomic bacterial artificial chromosome （BAC） libraries of Zhikong scallop Chlamys farreri were constructed to promote our genetic and genomic research. High-quality megabase-sized DNA was isolated from the adductor muscle of the scallop and partially digested by BamH I and Mbo I, respectively. The BamH I library consisted of 53 760 clones while the Mbo I library consisted of 7 680clones. Approximately 96 % of the clones in BamH I library contained nuclear DNA inserts in average size of 100 kb, providing a coverage of 5.3 haploid genome equivalents. Similarly, the Mbo I library with an average insert of 145 kb and no insert-empty clones, thus providing a genome coverage of 1.1 haploid genome equivalents.     

海鞘血细胞培养和细菌污染控制

以海鞘(Ciona savignyi)为研究对象,考察了海鞘血细胞原代培养方法及培养中细菌污染的鉴定和控制.血细胞在培养1 h后贴壁,细胞基本以圆形细胞为主,变形细胞一般呈不规则形.变形细胞存活时间较短,而圆形细胞存活时间较长.对于培养过程中的细菌污染,通过细菌分离、培养和纯培养发现两类菌株检出率较高,均为革兰氏阴性菌.经PCR扩增16S rDNA基因序列片断,结果显示这两类菌株分别属于弧菌属(Vibrio)和施万氏菌属(Shewanella).药敏试验结果显示,弧菌属对氯霉素和环丙沙星等具有较强的敏感性;对施万氏菌属较敏感的药物依次为亚胺培南和氯霉素等.最后比较了几种抗生素组合控制血细胞培养中细菌污染的使用效果,其中氯霉素、亚胺培南加双抗的组合有较好的抑菌效果并对培养细胞的贴壁和生长没有影响.     

凡纳滨对虾C-型凝集素LvLec2 对不同刺激的免疫应答

克隆获得了与中国明对虾(Fenneropenaeus chinensis)C-type lectin 3同源的C-型凝集素LvLec2基因序列,并通过Real-time PCR研究了其在脂多糖(lipopolysaccharide,简称LPS)、灭活溶壁微球菌(Micrococcus lysodeikticus)和白斑综合症病毒(white spot syndrome virus,简称WSSV)刺激后的转录表达变化。结果表明,LvLec2编码157个氨基酸,含有1个糖识别结构域(carbohydrate recognition domain,简称CRD),其CRD结构域中具有决定糖结合特异性的基序"EPS"(Glu118-Pro119-Ser120)序列。系统进化树分析表明LvLec2与已发表的凡纳滨对虾(Litopenaeus vannamei)C-型凝集素亲缘较远。LPS、灭活溶壁微球菌和WSSV刺激后,LvLec2基因在肝胰脏中的转录表达有明显的变化但表达模式不同。LvLec2可能作为模式识别受体参与了对虾对病原的识别或防御。     

微卫星筛选及其在对虾遗传学研究中的应用

Litt等[1]1989年在心肌肌动蛋白的基因内扩增了一种2核苷酸重复并创造了"微卫"(microsatellite)这个名词.后来,Edward等[2]称微卫星为简单重复序列(simple sequence repeat,SSR).现在人们一般把微卫星定义为1～6 bp 基元组成的重复序列.     

日本对虾野生种群和养殖种群遗传结构的RAPD标记研究

于１９９７年４月在福建和山东青岛分别采集地虾野生种群和养殖群体的样品,采用ＲＡＰＤ标记技术对其遗传结构进行初步研究。用２０个随机引物在野生种群和养殖群体中分别扩增出１５７和１５３条ＤＮＡ片段,其中多态性段数分别为８５和５８条,多态性片段的比例分别为５４．１４％和３７．９１％,平均杂合度分别为０．２５１７和０．１３４３。     

暴发性流行病病原对中国对虾越冬亲虾人工感染的研究

从典型的白斑症病虾中,分离出一种杆状病毒,属对虾白斑病杆状病毒［３］。为了探讨对虾暴发性流行病的发病机理及传播机制,作者于１９９６～１９９７年冬季,采用青岛地区当年夏季暴发典型白斑症的病虾,对中国对虾健康的越冬亲虾进行了白斑病杆状病毒的感染试验。１　材料和方法１．１　病虾标本与病原分离具典型白斑症状的病虾,于１９９６年８月取自青岛城阳区红岛邵格庄发病的虾场,保存于－７０℃。剥离病虾的胃、肠及鳃组织约１０ｇ,加液氮充分研磨,加９０ｍｌ０．０１ｍｏｌ／ＬＰＢＳ（０．２ｍｏｌ／ＬＮａ_２ＨＰＯ_４－Ｎａ…     

凡纳滨对虾P23蛋白基因的筛选、表达和生物信息学分析

克隆凡纳滨对虾极端体重个体的组织差异表达基因P23基因并进行生物信息学分析, 为进一步研究该基因的功能以及凡纳滨对虾的分子选育等研究提供信息。以凡纳滨对虾雌虾极端体重个体腹部肌肉为实验材料, 利用抑制消减杂交(SSH)技术构建极大体重雌性个体和极小体重雌性个体腹部肌肉组织正反向消减cDNA文库:正库(以极大体重个体为试验组, 以极小体重个体为驱动组, L-S)和反库(以极小体重个体为试验组, 以极大体重个体为驱动组, S-L)消减cDNA文库, 并采用实时定量技术分析P23基因的组织表达规律, 利用生物信息学对其功能和结构进行预测。以β-actin为看家基因检测两个文库的消减效率分别为210和25, 实时定量技术分析结果表明P23基因在体重的调节中起上调作用。P23基因编码区序列全长495bp, 编码165个氨基酸, GenBank登录号:JF806619。生物信息学分析发现P23蛋白部分序列含有强疏水性, 无跨膜螺旋结构, 两种信号肽预测都显示P23含有信号肽。     

Rapid, sensitive detection of Vibrio anguillarum using loop-mediated isothermal amplification

Vibrio anguillarum is an important bacterial pathogen of aquatic organisms and a significant problem in aquatic farming. The rapid detection and identification of V. anguillarum, and other pathogens that infect marine organisms, is crucial to effective disease management. In this study, we developed a loop-mediated amplification (LAMP) assay to detect V. anguillarum in an hour in a single tube without the need for thermal cycling. Conserved regions of the metalloproteinase (empA) gene of V. anguillarum served as the targets for primer design. A fragment of the empA gene was amplified at 65°C in the presence of the primer mixture and Bst DNA polymerase. In the optimized LAMP assay, 6.7 pg of V. anguillarum DNA could be detected. Six strains of V. anguillarum and 17 strains of non-V. anguillarum bacteria were used in this study to evaluate the species specificity of the primers. The six V. anguillarum strains gave a positive result in the LAMP assay. This method was also validated in V. anguillarum-infected fish. This LAMP method is more sensitive than PCR in the detection of V. anguillarum and shows good species specificity. The LAMP assay is therefore an effective method for the quick detection of V. anguillarum both in the laboratory and in the field.     

Heat-shock protein 70 expression in shrimp Fenneropenaeus chinensis during thermal and immune-challenged stress

Using western immunoblotting, we obtained heat-shock protein 70 (HSP70) induction data and distribution in different tissues from shrimp Fenneropenaeus chinensis during thermal and immune-challenged stresses. This is probably the first report of the effects of various stressors on the expression of HSP70 in shrimp.HSP70 was prominently induced in hepatopancreas and gills, but not in muscle, eyestalk and hemolymph, when the shrimp were exposed to heat shock and Vibrio anguillavium-challenged stresses. Cold shock and WSSV treatment had no significant effects on the levels of HSP70 expression in all tissues examined. HSP70 induction was greatest after 2 h exposure to heat shock stress, which was elevated after acute heat shock exposure of 10℃ above ambient temperature.     

Chromosome Behavior of Heat Shock Induced Triploid in Fenneropenaeus Chinensis

INTRODUCTIONTriploidanimalsareusefulbecausetheyaresterile ,characterizedbypositivegrowthinthere productiveseason ,usefulfortesting ,forintroductionofnon nativespecies ,andforprotectionofde velopedstrains.Theinductionoftriploidyhadbeenreportedinmanyaquaculturespecies .Triploidshavebeensuccessfullyinducedinshrimpusingtemperatureshockorchemicalshock (CB ,6 DMAP)in 4species :Sicyoniaingentis (Xiangetal.,1 991 ) ,Fenneropenaeuschinensis (Xiangetal.,1 992 ;Daietal.,1 993;Baoetal.,1 993;Li…