Identification and Characterization of Cell Wall Associated Silica Affinity Protein of Phaeodactylum tricornutum

• Main Authors: Chien, Ya-Yun, 錢亞筠
• Other Authors: Lin, Han-Jia
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/745pn6

碩士 === 國立臺灣海洋大學 === 生命科學暨生物科技學系 === 105 === Diatoms, a kind of single-celled algae, are the dominant phytoplanktons in the ocean. Each diatom has a cell wall, also known as frustule, which is made of biosilica (SiO2) and bearing the species-specific pattern. Based on the pattern of fructule, diatoms may be denoted as Coscinodiscophyceae and Pennales. Currently, the study of fructule formation was focused on Coscinodiscophyceae, especially in Thalassiosira pseudonana. Some proteins isolated from fructule are identified as silica affinity proteins, which can convert and deposit silicate(aq) into silica(s), including soluble silaffin and insoluble cingulin. However, sequence homologous proteins from was not identified in Pennales, such as Phaeodactylum tricornutum. In this study, we screened the genome database of P. tricornutum and found several candidate proteins with unusual high percentage of lysine and serine residues that is one of the characteristic of silica affinity protein. Next, these candidates were cloned and recombinant with a green fluorescent protein (EGFP) gene in a transgenic vector, respectively. After transfection and screening, transgenic P. tricornutum were observed under a fluorescence microscope to identify the cellular location of these candidate genes. We also isolated frustule of trangenic P. tricornutum to make sure which fusion protein was located in frustule. In addition, the gene expression patterns were also examed when P. tricornutum cultured in different silicate condition. Though our investigation, one silaffin-like protein in P. tricornutum was identified. We are going to further characterize this protein and use it to study the frustule formation mechanism in Pennales.