Bao Zhenmin

The changes of three components in coelomic fluid ofUrechis unicinctus exposed to different concentrations of sulfide

The changes in heme (associated with hemoglobin), hemoglobin and hematin in the coelomic fluid of marine worm,Urechis unicinctus, exposed to different concentrations of sulfide, were investigated using biochemical techniques. When exposed to different sulfide concentrations for up to 96 h, the relative amounts of the three components changed in a regular pattern suggesting that the coelomocytes play an important role in the worms tolerance to sulfide. The possible roles of heme compounds in sulfide tolerance of this species are discussed on the basis of our experimental data.

Genetic variation in two sea cucumber (Apostichopus japonicus) stocks revealed by ISSR markers

Sea cucumber Apostichopus japonicus samples were collected in Changdao, Penglai (PL), 27 individuals, and Lingshandao, Qingdao (QD), 30 individuals, in the Shandong Peninsula, China. Ten SSR primers were used to assess the genetic variation and relationship between and within the two stocks. Respectively, for each stock, the percentage of polymorphic bands was 85.2% and 86.9%; the gene diversity was 0.360 5 and 0.342 8; and the Shannon’s information index was 0.515 0 and 0.499 0. At species level, the percentage of polymorphic bands was 92.2%, the total gene diversity was 0.378 9 and the Shannon’s information index was 0.550 4. The coefficient of overall genetic differentiation and the genetic distances between the stocks were also calculated to be 0.073 0 and 0.079 6 using the POPGENE program. Results show that the genetic diversity of the two stocks is still large but the genetic distance between the two stocks is close. A dendrogram was constructed for the 57 individuals from the two stocks, showing that the genetic structure was unitary for PL stock but complex for QD stock.

Effect of Epinephrine on the Settlement and Metamorphosis of Manila Clam Larvae

Chemical inducement and DDRT-PCR (differential display reverse transcription PCR) are adopted to investigate the effect of epinephrine (EPI) on the settlement and metamorphosis of Manila clam larvae. Chemical inducement shows that EPI has an effect to some extent on the metamorphosis of Manila clam larvae at all concentrations and in all treatments designed. The most significant result of inducement is obtained at the concentration of 10−6 mol L−1 and for 4h. DDRT-PCR using six primer pairs shows that the gene expression pattern is quite different between EPI treatment and the control. Three hundred and forty-three amplification bands are obtained in total, among which, 67 (19.53%) are differentially appeared. Therefore, EPI has an effect on the gene expression of the eye spot larval Manila clam. It can be hypothesized that EPI is a settlement and metamorphosis inducer for Manila clam. EPI may lead to larvae settlement and metamorphosis by binding to the receptors on the membrane and then changing the gene expression of larvae cells.

An Efficient Procedure for Isolating Microsatellite DNAs from Sea Cucumber (Apostichopus japonicus)

The construction of enrichment library proves to be one of the efficient approaches for isolating microsatellites in this study. The genomic DNA of sea cucumber was digested with HaeIII and size-selected DNA fragments (250–700 bp) were ligated to an adaptor. Microsatellite-containing sequences were captured by using a combination of GA and CA probes, which were attached to a nylon membrane. The microsatellite enrichment library constructed in this study consisted of approximately 700 clones. Two hundred and thirty-two clones reacted positively after the library screening procedure. Of the 50 clones sequenced, all contained at least one microsatellite and one duplicate clone was found. Approximately 86% of the sequenced fragments permitted to design primers for sequence tagged microsatellite site (STMS).

Existence of microsatellites in expressed sequence tags of common carp (Cyprinus carpio L.) available in GenBank dbEST database

Common carp expressed sequence tags (ESTs) were analyzed for the existence of microsatellites, or simple sequence repeats (SSRs). In the NCBI dbEST database, a total of 10612 sequences were registered before December 31, 2004. A complete search of 2–6 nucleotide microsatellites resulted in the identification of 513 SSR-containing ESTs, accounting for 4.8% of the total. Cluster analysis indicated that 73 sequences of SSR-containing ESTs fell into 27 groups and the remaining 440 ESTs were indenpendent. A total of 467 unique SSR-containing ESTs were identified. These EST-SSRs contained a variety of simple sequence types, and di- and tri-nucleotide repeats were the most abundant, accounting for 42.1% and 27.9% of the whole, respectively. Of the dinucleotide repeats, CA/TG was the most abundant, followed by GA/TC. BLASTx search showed that 38.1% of the SSR loci could be associated with genes or proteins of known or unknown function. BLASTx searches of SSR-containing ESTs also showed high frequencies (98/179) of hits on zebrafish sequences.

基于16S rDNA基因序列探讨引进物种紫扇贝( Argopecten purpuratus )在海湾扇贝属( Argopecten )中的分类地位

秘鲁紫扇贝（Argopecten purpuratus）于2008年被引进我国，与海湾扇贝（Argopecten irradians irradians）成功杂交。为探讨紫扇贝在海湾扇贝属中的分类地位及与相似种海湾扇贝的进化关系，随机选取紫扇贝和海湾扇贝各10个个体，利用通用引物进行16S rDNA基因片段扩增并双向测序。序列分析结果表明，紫扇贝和海湾扇贝扩增获得基因片段长度均为542bp，碱基组成A+T所占比例分别为紫扇贝54.2%和海湾扇贝55.1%。紫扇贝和海湾扇贝种内分别有6和10个变异位点，种间有41个变异位点，且种内和种间变异均为转换或者颠换，无插入/缺失类型。结合GenBank同源序列信息，对海湾扇贝属物种进行遗传距离分析及分子系统树的构建。结果表明：海湾扇贝属7种扇贝[紫扇贝（A.purpuratus）、A.irradians、墨西哥湾扇贝（A.i.concentricus）、A.i.irradians、A.nucleus、A.gibbus和A.ventricosus]在系统树上聚为2支，与紫扇贝亲缘关系最近的是A.ventricosus，且该两种扇贝单独聚为一支。其余5种扇贝中，A.irradians、A.i.concentricus和A.i.irradians最先聚为一支，支持A.i.concentricus和A.i.irradians为A.irradians两个亚种的结论；另外，A.nucleus与以上三者也有很近的亲缘关系。该研究结果为海湾扇贝属物种进化、迁移及其遗传育种研究提供了理论基础。