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生物晶片 DNA 微陣列實驗 (DNA Microarray)

生物晶片 DNA 微陣列實驗 (DNA Microarray). Introduction. Molecular biology and central dogma DNA, RNA and protein Replication, transcription and translation Those basic concepts would help you understand the design of this experiment Microarray RNA expression analysis High throughput. DNA & RNA.

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生物晶片 DNA 微陣列實驗 (DNA Microarray)

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  1. 生物晶片DNA微陣列實驗 (DNAMicroarray)

  2. Introduction • Molecular biology and central dogma • DNA, RNA and protein • Replication, transcription and translation • Those basic concepts would help you understand the design of this experiment • Microarray • RNA expression analysis • High throughput

  3. DNA & RNA • Nucleotides (base, suger, phosphate) • Base – DNA: A-T, C-G RNA: A-U, C-G Double Helix structure

  4. Transcription Translation RNA (mRNA) DNA Protein cDNA Central dogma

  5. Protein synthesis • Codon (genetic code) Three-nucleotide sequences used in mRNA to specify one of the 20 amino acid used for protein synthesis

  6. Amino Acid

  7. 基因小常識 • 人類DNA總長度大約是30億個鹼基對(base pairs) • 生物體內有多少個基因? 人類約二萬至二萬五千個 (1%~1.5%) 蛔蟲約一萬九千個 酵母菌約六千個 結核病菌約四千個 • 不同個體間的DNA差異有多大? 人與人之間，差異約0.2% 人與黑猩猩之間，差異約2% 我們尚不了解其作用的DNA，約佔97% By Prof. Eric Y. Chuang

  8. Northern Blot (北方墨點法) • 分析該基因轉錄出的mRNA大小、序列、數量

  9. Polymerase Chain Reaction (PCR) DNA

  10. Polymerase Chain Reaction (PCR) • DNA Amplification http://www.sciencemedia.com/website/demos/biochem/PCRMutagenesis.html

  11. Microarray • Isolation of RNA • Salt, DNA, polysaccharide, and other contaminants • Decrease the background noise • Labeling and hybridization • As markers for target sequences • mRNA reverse transcription to cDNA and hybridization with the microarray chip • Scan and data analysis • Analysis of fluorescence intensity • Choose candidates

  12. Microarray http://www.youtube.com/watch?v=SNbt--d14P4&feature=related

  13. Cy5 Cy3

  14. Profiling Gene Expression • Clinical research -- tumor classification -- tumor vs. normal tissue • Basic research -- treated vs. untreated -- wild-type vs. mutant/knockout By Prof. Eric Y. Chuang

  15. Experiment planning • 預備實驗 • 微量吸管(Pipetman) 操作 • 正式實驗 • 時間：12/8 全天＆ 12/9 上午 --- 3、4、5、7、8 12/15 全天＆ 12/16 上午 --- 1、2、6、9、10 • 地點：長興街生技中心 莊曜宇老師實驗室(男七舍對面) 吳逢昌 助教

  16. 正式實驗 • 請注意實驗講義中的操作步驟與注意事項 • 使用tube 放入離心機或者進行均勻混合時，請將蓋子蓋緊 • 藥品加入的步驟請依序處理 • 反應時間請確實掌握 • 任何步驟的操作精確與否都會影響最後的實驗結果 • Everything is new, please be serious

  17. Artifact

  18. 實驗報告 • 實驗報告中至少包括： • 將你的實驗操作步驟畫成流程圖 • 回答實驗問題

  19. 預備實驗 • Pipetman 操作實習 • 不同型號的pipetman 操作的注意事項 • Pipetman 量測微量液體的精準度 • 生化實驗中幾乎不能沒有它

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