传染病病原体的诊断　课件教材教学课件.pptVIP

High Sensitivity Scanner 0.05 CPSM CPSM - what does it mean? Chromphores per square micron 0.05 cpsm or 5 molecules of dye per 10 micron pixel. Competitors best is 10 molecules of dye per 10 micron pixel. To achieve high sensitivity: Dynamic Autofocus Laser stability Control High resolution scanning 0.05 CPSM Detect 5 dye molecules in 10um pixel 0.1 CPSM Detect 10 dye molecules in 10um pixel eArray Delivering Custom Content and Formats Back Name Your Design Select Array Format Prokaryotic Array Custom Designs Agrobacterium Anopheles (AG) and Plasmodium (PB)...ie. malaria microarray Archaeon, Halobacterium sp. Bacillus cereus Bacterial pathogens Bifidobacterium longum Bordetella Pertussis Chlamydophila abortus Culex pipiens Cyanobacterium Synechocystis E.Coli Erwinia carotovora atroseptica/Solanum tuberosum Lactobacillus plantarum Methylobacterium extorquens AM1 Plasmodium falciparum Pseudomonas aeruginosa S.aureus; N.meningococcus; E.coli Salmonella typhimurium Staphylococcus aureus SARS Coronavirus Human Coronavirus OC43 Human Coronavirus 229E GreeneChip 1.1 CATCTGGTGTGGAAGCCTTATCGGAACAAGGACCAGAGCCACCTGGGCTGAGAACATCTA Sample: WNV NY99 100,000 RNA copies/ml 32/45 top hits are WNV-specific; remainder are JEV group or other flavivirus-genus probes Collaboration between the Greene Laboratory of Columbia University, ICTV , Northeast Biodefense Center, and Agilent Corporation CATCTGGTGTGGAAGCCTTATCGGAACAAGGACCAGAGCCACCTGGGCTGAGAACATCTA Sample: SARS-CoV 100,000 RNA copies/ml GreeneChip 1.1 Columbia (Briese, Lipkin, Liu, Lussier Palacios), ICTV (Büchen-Osmond), and Agilent (Hirschberg) 251,000 sequence database, 1710 vertebrate viruses 未知新病原体的发现 发现新病毒的主要技术路线 表达cDNA文库筛选技术 宽范围PCR（基于保守序列的PCR） 差异显示技术： 代表性差异分析（RDA） 抑制性消减杂交（SSH） 序列非依赖的扩增： 随机PCR（randome PCR） 序列非依赖的单引物扩增（SISPA） cDNA 文库筛选 cDNA文库筛选常被用来检出RNA病毒的核酸。 收集病毒颗粒（感染组织匀浆过滤，或者血浆超速离心以收集病毒颗粒），提取RNA，逆转录成c DNA ,构成c DNA表达文库。 最常用的表达型载体是λgt11,插入的c DNA片断可以融合蛋白形式表达，保留抗原性。