Synchronized Culture Of Skeletonema Dohrnii And Screening Of Silaffin Genes

As a group of single-celled eukaryotic algae and very easily been found in waters all over the world,diatoms account for 20%of the global annual carbon productions.Their cosmopolitan distribution make them vital in aquatic food webs and biogeochemical cycles such as the carbon cycle in the ocean.The unique silica cell walls(frustule)of diatoms provide mechanical protection for the algae cells,furthermore,help on their thriving successfuly.The ability of diatoms to make silica cell walls has been a subject of fascination for centuries.Nowdays,the study on the silicification of diatom cell walls involves a multi-disciplines,such as,microscopy,chemistry,biochemistry,material characterization,molecular biology,omics,transgene,and others.Skeletonema is a type of central diatom that is widely distributed in oceans all over the world.The experimental algae species in this thesis is Skeletonema dohrnii,which is a common bloom species in coastal waters.Through the method of cell synchronization and microscopic observation,this paper records and discusses the growth status of S.dohrnii under normal and silicon-depletion conditions.In addition,this paper also discusses the application of the python program in the screening of silaffin-like proteins.1.The light,temperature,and nutrients in environmental waters play crtical role on the growth of diatoms.The formation of the diatom cell wall involves many biosynthesis processes:genes,protein products,exocytosis,cell cycle,cytoskeleton,etc.Depletion of silicate in the medium will not only prevents the formation of diatom cell walls,and changes the expression of silicon-related proteins,but also inhibits the division of S.dohrnii cells,keeps most of the cells at the same phase of the cell cycle.In this thesis,most of the algae cells stayed in the G2+M phase by silicon depletion treatment of S.dohrnii.2.The method of microscopic observation is the main method for detecting cell morphology of diatom cells and their growth status during synchronization.Ordinary light microscopy is the best way to observe the original morphology of S.dohrnii in the culture medium.Observation of the microstructure of S.dohrnii by electron microscopy is helpful for us to have a further understanding of the cell wall structure of S.dohrnii.Re-add silicon element to the S.dohrnii culture medium that the algal cells have been limited to the same period,[2-(4-pyridyl)-5-((4-(2-dimethylamino ethyl aminocarbonyl)methoxy)phenyl)oxazole](PDMPO)can be used to stain the neonatal cell wall.Observation of the stained algae cells with a fluorescent microscope will help us understand the process of cell wall formation of S.dohrnii.3.As a protein that is indispensable in the process of diatom siliceous wall formation,the non-conservation of silaffins is significant.It is the reason why its genes are difficult to identify and screen out from the genome data.In this thesis,6 silaffin mRNA sequences were screened from the differential transcriptome data of S.dohrnii by sequence alignment.Using python programs and characteristic motifs,29 silaffin-like sequences were screened out from the proteins that predicted by 5 diatom's genomes.Using python programs to screen non-conservative proteins with characteristic motifs may become a new method for screening and identifying proteins.