中国南部沿海尖头斜齿鲨遗传多样性研究

采用随机引物扩增多态性DNA(RAPD)技术,对中国南部沿海闽东霞浦-浙南瑞安海域(MD)、闽南-台湾浅滩渔场(MN)、粤西湛江-阳江海域(YX)、北部湾海域(BBW)4个尖头斜齿鲨地理群体遗传多样性进行分析,29个随机引物在4个群体中共检测出282个位点.4个群体各自检测出的位点数分别为273、274、272、276,其中多态位点数分别为54、57、52、45,多态位点比例分别为19.78%、20.80%、19.12%、16.30%.Shannon多样性指数分别为0.104、0.107、0.103、0.090,表明尖头斜齿鲨群体的遗传多样性水平较低.遗传距离和UPGMA聚类分析结果显示尖头斜齿鲨的基因交流模式属于距离隔离模式,遗传差异大小与地理距离远近相关.进行分子方差分析(AMOVA)得到遗传变异固定指数(Fst＝0.019,p＜0.05),显示大部分变异(98.06%)发生在群体内部,群体间变异较小(1.94%).     

海带和长海带配子体无性繁殖系微卫星DNA多态性比较分析

探讨促进海带配子体克隆资源的有效保存和合理利用途径,进行海带良种和杂交海带培育,开发了18个海带微卫星DNA标记,并用这些标记分析了海带和长海带配子体克隆的遗传多样性.这些标记相互独立,至少在1个物种中呈多态性.在36份海带配子体中,这些标记检测到的等位基因数量从2～7个不等,揭示的基因多样性在0.21(H144)和0.75(H120)之间,揭示的香农氏信息指数在0.37(H144)和1.55(H120)之间.长海带和海带的平均等位基因数(1.5/3.3),平均基因多样性(0.12/0.45)和平均香农氏信息指数(0.20/0.81)显著不同.根据总样本确定的前2个最丰富等位基因中,绝大多数的频率在长海带和海带之间存在显著差异.长海带和海带之间在基因多样性和香农氏信息指数反映的遗传多样性上的平均遗传分化分别达35.6%和36.9%.     

Inheritance mode of microsatellite loci and their use for kinship analysis in the Pacific oyster （Crassostrea gigas）

Five full-sib families of the Pacific oyster(Crassostrea gigas) larvae were used to study the mode of inheritance at eight microsatellite loci,and the feasibility of these markers for kinship estimate was also examined.All eight microsatellite loci were compatible with Mendelian inheritance.Neither evidence of sex-linked barriers to transmission nor evidence of major barriers to fertilization between gametes from the parents was shown.Three of the eight loci showed the presence of null alleles in four families,demonstrating the need to conduct comprehensive species-specific inheritance studies for microsatellite loci used in population genetic studies.Although the null allele heterozygotes were considered as homozygotes in the calculation of genetic distance,offspring from five full-sib families were unambiguously discriminated in the neighbor-joining dendrogram.This result indicates that the microsatellite markers may be capable of discriminating between related and unrelated oyster larvae in the absence of pedigree information,and is applicable to the investigation of the effective number of parents contributing to the hatchery population of the Pacific oyster.     

Characterization of defensin gene from abalone <Emphasis Type="Italic">Haliotis discus hannai</Emphasis> and its deduced protein

Defensin is one of preserved ancient host defensive materials formed in biological evolution. As a regulator and effector molecule, it is very important in animals＇ acquired immune system. This paper reports the defensin gene from the mixed liver and kidney cDNA library of abalone Haliotis discus hannai Ino. Sequence analysis shows that the gene sequence of full-length cDNA encodes 42 mature peptides （including six Cys）, molecular weight of 4 323 Da, and pI of 8.02. Amino acid sequence homology analysis shows that the peptides are highly similar （70% in common） to other insects defensin. Because of a typical insect-defensin structural character of mature peptide in the secondary structure, the polypeptide named Haliotis discus defensin （hd-def）, a novel of antimicrobial peptides, belongs to insects defensin subfamily. The RT-PCR result of Haliotis discus defensin shows that the gene can be expressed only in the hepatopancreas by Gram-negative and positive bacteria stimulation, which is ascribed to inducible expression. Therefore, it is revealed that the Haliotis discus defensin gene expression was related to the antibacterial infection of Haliotis discus hannai Ino.     

Inheritance mode of microsatellite loci and their use for kinship analysis in the Pacific oyster （Crassostrea gigas）

Five full-sib families of the Pacific oyster （Crassostrea gigas） larvae were used to study the mode of inheritance at eight microsatellite loci, and the feasibility of these markers for kinship estimate was also examined. All eight microsatellite loci were compatible with Mendelian inheritance. Neither evidence of sex-linked barriers to transmission nor evidence of major barriers to fertilization between gametes from the parents was shown. Three of the eight loci showed the presence of null alleles in four families, demonstrating the need to conduct comprehensive species-specific inheritance studies for microsatellite loci used in population genetic studies. Although the null allele heterozygotes were considered as homozygotes in the calculation of genetic distance, offspring from five full-sib families were unambiguously discriminated in the neighbor-joining dendrogram. This result indicates that the microsatellite markers may be capable of discriminating between related and unrelated oyster larvae in the absence of pedigree information, and is applicable to the investigation of the effective number of parents contributing to the hatchery population of the Pacific oyster.     

中国对虾(Fenneropenaeus chinensis)“黄海1号”部分生长相关性状的QTL定位分析

在已构建好的中国对虾"黄海1号"F2代遗传连锁图谱上,使用WinQTLcart2.5软件进行复合区间作图定位,通过置换实验(1000次重复)确定连锁群显著性水平阈值。在对体重、各腹节长和尾节长8个性状的QTL定位分析中,共检测到29个QTL,其中与体重相关的QTL(BW1.2)、腹节1长相关的QTL(A1L2.1、A1L9.2、A1L17.3、A1L26.4)、腹节3长相关的QTL(A3L16.1)、腹节4长相关的QTL(A4L26.1)、腹节6长相关的QTL(A6L20.3)及尾节长相关的QTL(T3L17.3)等9个达到连锁群显著水平(P=0.05),对性状的贡献率为11.14%—23.60%,其中位于第16连锁群(LG16)上的与腹节3长相关的QTL(A3L16.1)的贡献率为23.60%,达到了极显著水平(P0.001),但它们的加性方向并不一样,除了与腹节1长相关的QTL(A1L2.1)和与腹节6长相关的QTL(A6L20.3)方向为负以外,其余7个主效QTL均呈正方向。本研究同时发现了与QTL紧密连锁的分子标记,如位于第1连锁群(LG1)上的分子标记D9f446、位于第17连锁群(LG17)上的M10f246、位于第26连锁群(LG26)上的SX18和位于第30连锁群(LG30)上的B2f260等,这些与QTL距离仅为0.01cM(厘摩)的分子标记,将为QTL精细定位提供参考。     

两种蛤仔群体遗传多样性的形态参数及AFLP分析

采用多变量形态度量学方法,对大连(DL)、青岛(QD)、厦门(XM)、珠海(ZH)4个菲律宾蛤仔群体和湛江(RV)1个杂色蛤仔群体的形态变异进行了比较研究,建立了群体形态聚类图和形态判别函数,结果较客观地显示了4个菲律宾蛤仔群体之间以及与杂色蛤仔群体之间的差异。利用4对引物组合对菲律宾蛤仔青岛群体与杂色蛤仔湛江群体共48个个体进行了AFLP分析,共得到216个位点,其中特有位点29个,可作为2个物种鉴别的分子标记。两种蛤仔都显示了较高的群体遗传多样性,菲律宾蛤仔群体内遗传相似度为0.6051,杂色蛤仔为0.6882,群体间的平均遗传相似度只有0.2968。群体内香农多样性指数菲律宾蛤仔为0.2526,略高于杂色蛤仔的0.2154。对所有个体聚类分析,同种个体可以很好地聚在一起,没有出现种间的交叉,表明两种蛤仔具有明显的遗传差异。     

海蜇养殖群体及自然捕获群体ITS 序列遗传分析

为调查海蜇(Rhopilema esculentum)自然海区群体和养殖群体遗传多样性,对烟台莱州湾和江苏海州湾自然海区捕获群体及威海养殖群体24个个体的ITS序列进行了PCR扩增和序列分析,获得DNA片段长度在1 080~1 096 bp之间,包括完整的ITS1,5.8S和完整ITS2序列。结果表明,在所测碱基序列中共发现26个变异位点,4处多碱基插入位置,29个插入缺失位点。群体内平均核苷酸差异数K和平均核苷酸多样性指数Pi为2.179~2.750和0.002 02~0.002 54,群体间平均核苷酸差异数K值波动在2.797~3.031之间,遗传距离在0.002 30~0.002 81之间,遗传分化指数(Fst)值在0.032 50~0.730 8之间,群体内及群体间遗传多样性指标数值比较相近,群体间的遗传距离比较小,群体遗传结构相似,群体间没有明显的遗传分化。     

大珠母贝微卫星DNA标记的分离与筛选

采用生物素标记的(CA)15探针和磁珠富集法构建了大珠母贝(Pinctada maxima)微卫星DNA文库,随机挑选1200个菌落,经PCR筛选得到298个候选克隆,成功测序246个,分析获得251个微卫星序列,其中完美型、非完美型和混合型分别占63%、32%和5%。除探针中使用的CA重复外,还得到AT、GT、TC、AG、GC、TAA、CAA、AGG、GATA、GACA、GTGC、CGTC、GACG、CTGT等重复序列。设计引物90对,挑选其中30对合成并筛选出21对能在大珠母贝基因组有效扩增的引物。种群PCR扩增后利用变性聚丙烯酰胺凝胶电泳法进行检测,获得了10个多态位点,共检测出59个等位基因,片段长度范围为133~444 bp。各位点等位基因数2~8个,平均等位基因数5.9个;有效等位基因数为1.342 3~6.000 0,平均为4.124 0;多态性信息含量(CPI)为0.222 5~0.811 8,平均0.7179;期望杂合度(He)为0.259 3~0.847 5,平均0.717 9;观测杂合度(Ho)为0.300 0~0.800 0,平均0.533 3。这些微卫星多态性标记的获得,为进一步开展大珠母贝遗传育种、保护生物学和种群遗传多样性等研究奠定了一定基础。     

基于线粒体DNA部分片段探讨石鲽与星突江鲽的亲缘关系

比较分析了石鲽和星突江鲽线粒体基因组COI、Cytb基因以及D-loop片段总长度为1175 bp的核苷酸序列,探讨了石鲽与星突江鲽的亲缘关系。2种间共检测到95处核苷酸替代,蛋白质编码基因上的核苷酸替代主要是第三密码子位点上的同义替换。核苷酸组成分析结果表明:3个目的片段的鸟嘌呤(G)含量普遍较低,在2个蛋白质编码基因第三密码子位点上表现得尤为明显。线粒体COI、Cytb基因和D-loop片段序列分析显示石鲽与星突江鲽间平均遗传距离分别为0.043、0.062和0.153,属于属内种间差异水平。2种鲽鱼在线粒体基因组不同片段上存在明显的遗传分化,核苷酸替代速率由大到小依次为D-loopCytbCOI。贝叶斯法、邻近距离法和最大简约法构建的系统发育树一致,皆显示石鲽与星突江鲽遗传关系很近。基于Cytb基因片段序列的分析结果表明:石鲽与星突江鲽的分歧时间约为185万年,分化事件发生于更新世中期(Middle Pleistocene)。