【摘要】 链状亚历山大藻是赤潮爆发的重要原因种。本文拟构建与高磷浓度诱导相关的链状亚历山大藻基因表达抑制性消减文库,并在锰条件诱导生长细胞中进行基因表达的验证,为获得赤潮爆发性生长相关基因及其应用奠定基础。构建了不同磷浓度条件下链状亚历山大藻的消减文库,挑取383个克隆,经检测有348个为阳性克隆,阳性率达90.9%,后采用2轮斑点杂交,即将获得的348个克隆的PCR产物纯化后先与低磷和高磷的cDNA探针进行杂交,选取杂交信号差异在1.4倍以上的克隆,再分别与无锰及高锰的cDNA探针杂交,选取杂交信号差异在1.4倍以上阳性克隆12个测序,获得了6个可分析的基因片段,分别与生长、翻译起始和能量传递有关,其中2个为编码未知功能蛋白基因片段,1个与转化蛋白功能有相似性,但相似性较低。初步认为在赤潮爆发性生长中这些基因通过影响和加强转录调节、翻译的起始及能量传递等,起始或加速了细胞的快速生长分裂。更多还原

【Abstract】 Alexandrium catenella is an important causative dinoflagellate of harmful algal blooms(HABs).In the present paper,suppressive subtractive hybridization(SSH) library of the cells induced by high phosphor cultivating condition of Alexandrium catenella was constructed.The expression of the clones from the library were further demonstrated by the cells cultivated under the medium with manganese additive.The result will contribute to the obtaining of burst-growth related genes of HAB species and their applications.Suppressive subtractive library of Alexandrium catenella under different phosphor concentrations in cultivation medium were constructed after suppressive subtractive hybridization.Three hundred and eighty-three clones were obtained from the SSH library,involving 348 positive clones with the positive ratio of 90.9%.The PCR products from the 348 clones were purified,and then screened by two rounds of dot hybridization with the first round done by probes produced from the cDNA of cells cultivated under low and high phosphor conditions respectively.The clones showing a signal ratio higher than 1.4 between these two kinds of hybridization were selected and undergone a second round of dot hybridization which operated on the probes produced from the cDNA of cells cultivated under conditions with and without manganese additive,respectively.Twelve clones were obtained with a signal ration higher than 1.4 between these two kinds of hybridization and sequenced.Six gene fragmnents were identified with four of them showing the predicted function of growth,the initial of translation and energy delivering,respectively.The other two had unknown functions and one of which was weakly similar to transforming protein.It was suggested that the burst growth of this HAB species may relate to the influence or promoting of regulation of transcription,the initial of translation and energy delivering,thus initiating or accelerating the fast growth and division of the cells.更多还原