臺灣博碩士論文加值系統

Chlorella sorokiniana T89為台灣嘉義水稻田所篩選出的小球藻，其genomic DNA與cDNA之定序與註解至今仍未有研究發表。小球藻油脂含量高達30%，且富含不飽和脂肪酸，十分適合作為生質柴油或機能性食品。經培養不同生長狀態的小球藻，萃取其RNA並轉成cDNA，藉由Illumina 定序系統得到基因片段序列contigs，分析其表現量差異；進行NCBI與KEGG之BLAST進行註解，了解各contig所比對之基因與其參與的pathway map。從差異表現量的註解資料中，選取抗氧化機制中扮演重要角色之酵素─超氧歧化

The heat-talent Chlorella sorokiniana T89 was cloned from Chiayi paddy field. The genomic DNA and cDNA annotated data was unpublished so far. The lipid content of C. sorokiniana T89 was up to 30% and rich in unsaturated fatty acids which were suitable as biodiesel and functional foods. Culturs of C. sorokiniana T89 at different growth stages were used for RNA extracted and converted into cDNA for future sequencing analysis. Then get the contigs sequence data by Illumina sequencing system. Contigs were predicted their function via sequence alignment by using BLAST tools from NCBI and KEGG to find gene functions. Among the annotated genes with differential expression, the superoxide dismutase (SOD) which plays an importante role at antioxidant pathways was selected for further cloning, expression and biochemical characterization. In the future, we can use the annotation database to study the genes involved in physiological pathway of C. sorokiniana T89 andto developict as a good resource of biodiesel.

論文摘要 i
Abstract ii
圖目錄 vi
表目錄 viii
第一章 緒論 1
第二章 文獻回顧 2
2.1 小球藻Chlorella sorokiniana T89 2
2.2 核酸定序系統 3
2.3 BLAST (Basic Local Alignment Search Tool) 3
2.4 超氧歧化

1.Becker W: Microalgae in human and animal nutrition. Handbook of microalgal culture 2004:312–351.
2.Spolaore P, Joannis-Cassan C, Duran E, Isambert A: Commercial applications of microalgae. J Biosci Bioeng 2006, 101(2):87-96.
3.Certik M, Shimizu S: Biosynthesis and regulation of microbial polyunsaturated fatty acid production. J Biosci Bioeng 1999, 87(1):1-14.
4.Gill I, Valivety R: Polyunsaturated fatty acids, Part 1: Occurrence, biological activities and applications. Trends Biotechnol 1997, 15(10):401-409.
5.Kessler E: [Physiological and biochemical contributions to the taxonomy of the genus Chlorella. VI. Utilization of organic carbon compounds]. Arch Mikrobiol 1972, 85(2):153-158.
6.黃 檀, 林 榮: 比較耐熱性小球藻異營生長之特性. 師大學報：數理與科技類 2002, 47(1):31-40.
7.Sanger F, Nicklen S, Coulson AR: DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A 1977, 74(12):5463-5467.
8.Mardis ER: Next-generation DNA sequencing methods. Annu Rev Genomics Hum Genet 2008, 9:387-402.
9.Yamakura F, Sugio S, Hiraoka BY, Ohmori D, Yokota T: Pronounced conversion of the metal-specific activity of superoxide dismutase from Porphyromonas gingivalis by the mutation of a single amino acid (Gly155Thr) located apart from the active site. Biochemistry 2003, 42(36):10790-10799.
10.Corpas FJ, Barroso JB, del Rio LA: Peroxisomes as a source of reactive oxygen species and nitric oxide signal molecules in plant cells. Trends Plant Sci 2001, 6(4):145-150.
11.Corpas FJ, Fernandez-Ocana A, Carreras A, Valderrama R, Luque F, Esteban FJ, Rodriguez-Serrano M, Chaki M, Pedrajas JR, Sandalio LM et al: The expression of different superoxide dismutase forms is cell-type dependent in olive (Olea europaea L.) leaves. Plant Cell Physiol 2006, 47(7):984-994.
12.Abreu IA, Cabelli DE: Superoxide dismutases-a review of the metal-associated mechanistic variations. Biochim Biophys Acta 2010, 1804(2):263-274.
13.Campana F, Zervoudis S, Perdereau B, Gez E, Fourquet A, Badiu C, Tsakiris G, Koulaloglou S: Topical superoxide dismutase reduces post-irradiation breast cancer fibrosis. J Cell Mol Med 2004, 8(1):109-116.
14.Chothia C, Lesk AM: The relation between the divergence of sequence and structure in proteins. EMBO J 1986, 5(4):823-826.
15.Folch J, Lees M, Sloane Stanley GH: A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem 1957, 226(1):497-509.
16.Morrison WR: A Fast, Simple and Reliable Method for the Microdetermination of Phosphorus in Biological Materials. Anal Biochem 1964, 7:218-224.
17.Sun L, Chen M, Yang H, Wang T, Liu B, Shu C, Gardiner DM: Large scale gene expression profiling during intestine and body wall regeneration in the sea cucumber Apostichopus japonicus. Comp Biochem Physiol Part D Genomics Proteomics 2011, 6(2):195-205.
18.Beauchamp C, Fridovich, I.: Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Anal Biochem 1971, 44:276-287.
19.Huang JK, Wen L, Ma H, Huang ZX, Lin CT: Biochemical characterization of a cambialistic superoxide dismutase isozyme from diatom Thallassiosira weissflogii: cloning, expression, and enzyme stability. J Agric Food Chem 2005, 53(16):6319-6325.
20.Shen MY, Sali A: Statistical potential for assessment and prediction of protein structures. Protein Sci 2006, 15(11):2507-2524.
21.Zhou H, Zhou Y: Single-body residue-level knowledge-based energy score combined with sequence-profile and secondary structure information for fold recognition. Proteins 2004, 55(4):1005-1013.
22.Munoz IG, Moran JF, Becana M, Montoya G: The crystal structure of an eukaryotic iron superoxide dismutase suggests intersubunit cooperation during catalysis. Protein Sci 2005, 14(2):387-394.
23.Cuaresma M, Janssen M, Vilchez C, Wijffels RH: Productivity of Chlorella sorokiniana in a short light-path (SLP) panel photobioreactor under high irradiance. Biotechnol Bioeng 2009, 104(2):352-359.
24.Subramaniam R, Dufreche S, Zappi M, Bajpai R: Microbial lipids from renewable resources: production and characterization. J Ind Microbiol Biotechnol 2010, 37(12):1271-1287.
25.Dean AP, Sigee DC, Estrada B, Pittman JK: Using FTIR spectroscopy for rapid determination of lipid accumulation in response to nitrogen limitation in freshwater microalgae. Bioresour Technol 2010, 101(12):4499-4507.
26.Rodolfi L, Chini Zittelli G, Bassi N, Padovani G, Biondi N, Bonini G, Tredici MR: Microalgae for oil: strain selection, induction of lipid synthesis and outdoor mass cultivation in a low-cost photobioreactor. Biotechnol Bioeng 2009, 102(1):100-112.
27.Illman AM, Scragg AH, Shales SW: Increase in Chlorella strains calorific values when grown in low nitrogen medium. Enzyme Microb Technol 2000, 27(8):631-635.
28.Der JP, Barker MS, Wickett NJ, dePamphilis CW, Wolf PG: De novo characterization of the gametophyte transcriptome in bracken fern, Pteridium aquilinum. BMC Genomics 2011, 12:99.
29.Nies DH: Microbial heavy-metal resistance. Appl Microbiol Biotechnol 1999, 51(6):730-750.
30.Cleland WW: Dithiothreitol, a New Protective Reagent for Sh Groups. Biochemistry 1964, 3:480-482.
31.Grell L, Parkin C, Slatest L, Craig PA: EZ-Viz, a tool for simplifying molecular viewing in PyMOL. Biochem Mol Biol Educ 2006, 34(6):402-407.
32.Johnson M, Zaretskaya I, Raytselis Y, Merezhuk Y, McGinnis S, Madden TL: NCBI BLAST: a better web interface. Nucleic Acids Res 2008, 36(Web Server issue):W5-9.
33.Antonov AV, Dietmann S, Mewes HW: KEGG spider: interpretation of genomics data in the context of the global gene metabolic network. Genome Biol 2008, 9(12):R179.
34.Poirot O, O''Toole E, Notredame C: Tcoffee@igs: A web server for computing, evaluating and combining multiple sequence alignments. Nucleic Acids Res 2003, 31(13):3503-3506.
35.Prlic A, Bliven S, Rose PW, Bluhm WF, Bizon C, Godzik A, Bourne PE: Pre-calculated protein structure alignments at the RCSB PDB website. Bioinformatics 2010, 26(23):2983-2985.
36.Eswar N, Webb B, Marti-Renom MA, Madhusudhan MS, Eramian D, Shen MY, Pieper U, Sali A: Comparative protein structure modeling using MODELLER. Curr Protoc Protein Sci 2007, Chapter 2:Unit 2 9.
37.Kelley LA, Sternberg MJ: Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 2009, 4(3):363-371.
38.Chen CC, Hwang JK, Yang JM: (PS)2-v2: template-based protein structure prediction server. BMC Bioinformatics 2009, 10:366.
39.Wu S, Zhang Y: LOMETS: a local meta-threading-server for protein structure prediction. Nucleic Acids Res 2007, 35(10):3375-3382.