ABI公司测序仪的应用 Ph D Technical Product Specialist chenyd 22006 11 17 2006 Applied Biosystems 提要 IDNA测序仪应用概况测序仪应用概况 II基因突变快速扫描 基因突变快速扫描 SSCP III植物植物AFLP分析分析 IV细菌真菌鉴定 细菌真菌鉴定 MicroSeq ID V甲基化研究甲基化研究 VIBigDye XTerminator DNA测序仪应用概况 42006 11 17 2006 Applied Biosystems 310 3130 3130 xl3730 3730 xl ABI遗传分析仪家族 使用同样的试剂和软件使用同样的试剂和软件 1 4 16 48 96 52006 11 17 2006 Applied Biosystems 测序仪三大功能 DNA测序测序 测定测定DNA片段的碱基序列片段的碱基序列 片段分析片段分析 测定测定DNA片段的长度和颜色片段的长度和颜色 SNP分型分型 测定多态或点突变测定多态或点突变 62006 11 17 2006 Applied Biosystems 测序仪可以用来做什么 DNA测序测序 医学测序医学测序 细菌 真菌鉴定细菌 真菌鉴定 杂合子 突变检测杂合子 突变检测 SNP研究研究 SNaPshot Multiplex SNPlex 定量定量PCR TaqMan探针杂交探针杂交 片段分析片段分析 基于基于STR的法医鉴定 的法医鉴定 HID 基于基于STR的基因连锁定位 的基因连锁定位 LMS 突变筛查 突变筛查 SSCP 农牧业育种 农牧业育种 AFLP 72006 11 17 2006 Applied Biosystems 未知序列测序 100多种基因组已经 完成测序 许多新基 因组计划正在进行中 许多基因有待发现 理解生物学功能 解码人类疾病的路径 和机制 多种基因组已经 完成测序 许多新基 因组计划正在进行中 许多基因有待发现 理解生物学功能 解码人类疾病的路径 和机制 Diagram from Celera Genomics 82006 11 17 2006 Applied Biosystems 线粒体DNA测序 Deleterious mutations in gene coding regions Genetic variation in populations Human evolution Disease association studies 500 1000 copies of mtDNA per cell allows easier extraction MITOMAP A Human Mitochondrial Genome Database http www mitomap org 2004 Image provided by Molecular Probes photo gallery 2004 92006 11 17 2006 Applied Biosystems 比较基因组学 Genetic conservation and variation Finding answers to disease mechanisms How closely related are mice and humans How many genes are the same www pathology washington edu Mouse Karyotype ZooWeb Karyotypes home page Human Karyotype 人类 102006 11 17 2006 Applied Biosystems 医学测序 VariantSEQr Resequencing System A complete application solution for detecting variants in thousands of disease related human genes SeqScape Software v2 5 Allows automatic detection of heterozygotes Mutation Profiling Insertions Deletions Development of diagnostic prognostic assays for disease 112006 11 17 2006 Applied Biosystems 杂合子检测 发现新突变 122006 11 17 2006 Applied Biosystems 细菌和真菌鉴定 MicroSeq ID Analysis Software MicroSeq 500 16S Bacterial Identification Kit MicroSeq D2 LSU Fungal Identification Kit DNA Sequencing of the 16S ribosomal RNA gene Classification of Eubacteria Yeasts and Filamentous Fungi Sequence polymorphisms Bacterial category A biothreat agents Food pathogens Same day results 132006 11 17 2006 Applied Biosystems SAGE Analysis 基因组范围的基因表达图式分析 鉴定转录子 基因组范围的基因表达图式分析 鉴定转录子 mRNA 142006 11 17 2006 Applied Biosystems 基于测序的HLA配型 Donor Recipient compatibility prior to tissue transplantation HLA typing products distributed directly by Celera Diagnostics for research purposes only 1 866 235 3723 Referenced to GENBANK HLA C 6p21 3 major histocompatibility complex class I C http www ncbi nlm nih gov 152006 11 17 2006 Applied Biosystems 功能强大的片段分析 片段长度多态性 AFLP和T RFLP BAC指纹图谱 相对荧光定量 LOH和QMPSF 构象分析扫描突变 SSCP和CSCE InVivoScribe B细胞和T细胞clonality TILLING 162006 11 17 2006 Applied Biosystems 相对荧光定量 细胞淋巴瘤细胞淋巴瘤 cell lymphomas 及其他肿瘤中的等位基因丢失 染色体不平衡 部分和全部染色体丢失 非遗传的 及其他肿瘤中的等位基因丢失 染色体不平衡 部分和全部染色体丢失 非遗传的in vivo LOH LOH Loss of Heterozygosity P53 17p13 1 172006 11 17 2006 Applied Biosystems 寻找新的SNP位点 Mutation Profiling SNP Discovery and Analysis 182006 11 17 2006 Applied Biosystems SNP扫描 SNPlex Genotyping System SNaPShot Multiplex System SNP Validation Linkage Mapping and Association Studies Pharmacogenomic Research SSCP突变快速扫描 202006 11 17 2006 Applied Biosystems SSCP mobilitymobility rfurfu WTWT MUTMUT Wild TypeWild TypeMutantMutant A T G C A T G C Amplify Amplify region of region of interestinterest Heat denature in Heat denature in HiDiHiDi formamideformamide and and NaOHNaOH Rapidly chill and Rapidly chill and electrophoresis under electrophoresis under non denaturing non denaturing conditionsconditions ATGC 212006 11 17 2006 Applied Biosystems SSCP Workflow DNA Extraction TissueBloodCells Sample Setup Electrophoresis Sequence suspected mutants Bact DNA Amplification Post PCR Purification Data Analysis 222006 11 17 2006 Applied Biosystems SSCP data on ABI 3130 Series WT WT Mut1 WT Mut 2 Mut 1 Mut 2 植物AFLP 分析 242006 11 17 2006 Applied Biosystems AFLP技术原理 252006 11 17 2006 Applied Biosystems AFLP工作流程 Perform AFLP assay on genomic DNA templates Mix AFLP product with size standard and run on CE platform Analyze data using GeneMapper software Export Print Review final results Electropherograms Genotypes data in binary format 262006 11 17 2006 Applied Biosystems AB公司公司AFLP相关产品相关产品 Instruments 310 3130 3730 Size Standard GS500ROX Software GeneMapper v4 0 Kits Plant Mapping Kit Small Genomes 50 500Mb Regular Genomes 500 6000Mb Microbial Fingerprinting Kit Microbial and Fungal Genomes 272006 11 17 2006 Applied Biosystems 两个实例 Experiment 1 Maize genomic DNA templates Two parental samples P1 P2 and first generation offspring F1 from a cross between P1 and P2 Experiment 2 Hedysarum legume samples isolated from different locations in Utah AB plant mapping kit Optimal primers for maize but includes instructions for other plant systems Size standards GeneScan 500ROX Electrophoresis Standard 3