Chapter 18 - 204-325 1 Summary of the Last Lecture RFLP,...

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Unformatted text preview: 204-325 1 Summary of the Last Lecture RFLP, PCR and ASO hybridization are used to detect DNA polymorphisms. Almost all human beings are genetically distinct. SNP and the variable length of STRs or VNTRs can be used to identify an individual, detect paternity and for forensic applications. Genes can be cloned based on their genetic linkages with markers. Positional cloning is a powerful tool for identifying human disease genes. We are able to genetically modify or engineer many organisms for medical, agricultural and other purposes. Human gene therapy is limited to somatic tissues at present. 2 Today's Topics 1. Map-Based Genome Sequencing 2. Shotgun Genome Sequencing 3. Genome Projects and Analysis 4. Microarray 3 Topic 1 Map-Based Genome Sequencing 4 Steps in Genome Sequencing 5 Mapping Genome Genetic map based on meiotic recombination Cytogenetic map based on chromosome banding pattern Physical maps based on DNA analysis FISH map Radiation map Restriction map Clone contig map 6 FISH: Fluorescent In Situ Hybridization 7 Map Genes by Radiation Hybrid 8 Principle of Contig Map Contig: a set of overlapping segments of DNA : Sequence-Tagged Sites (STS) 9 Ordering YAC into Contig by STS 10 YAC Contig Map Assembled by STS Mapping 11 Map-Based Genome Sequencing Assemble within Contig 12 Topic 2 Shotgun Genome Sequencing 13 Shotgun Strategy 14 Shotgun Approach to Obtaining the Genomic DNA Sequence 15 Animation 1: Shotgun Sequencing 16 Topic 3 Genome Projects and Genome Analysis 17 Landmarks of Genome Projects Haemophilus Influenzae 1.8 Mb 1953 Double Helix 1977 DNA Sequencing 1986 Automated Sequencing 1995 First Genome 1996 Yeast 1997 E. coli 2000 2000 2003 1997 5Mb C. elegans Yeast Genome Drosophila Genome E. coli Arabidopsis Human 1998 1996 18 Tree of Life Branches with Sequenced Genome 19 Genome of Haemophilus Influenzae tRNA 1995 1.83 Mb rRNA 1743 ORF 20 E. coli Genome The Genome of E. coli K-12 4,639,221 base pairs 4288 Protein-coding Genes tRNA rRNA 21 Genome 0f E. coli 1997 22 A Close-Up View of a Portion of the E. coli Genome operons 23 Yeast Genome 1996 24 Yeast Proteome 6183 ORF 25 Arabidopsis Proteome 2000 120 Mb Genome 25,900 ORF 26 Human Genome: 3 Gb, ~23,000 ORF Draft Sequence April 14, 2003 Finished Human Genome: 99.999% accuracy, no gaps 27 Map-Based Sequencing of Human Genome 28 Comparison of the Two Approaches Map-based Shotgun Whole Genome Shotgun 29 Genome Sequencing Center 30 Features of Human Genome 31 Where to See the Human Genome? http://www.ncbi.nlm.nih.gov/genome/guide/human/ 32 Predicted Human Proteome 33 Synteny between Human and Mouse Genome Blocks of genes in humans are in closely linked blocks in the mouse genome. Such blocks of genes are said to be syntenic. 34 Synteny between Human and Mouse Genome Human Mouse Blocks of genes in humans are in closely linked blocks in the mouse genome. Such blocks of genes are said to be syntenic. 35 Features of Sequenced Genome 36 Gene Number Comparison 37 Genome Size, Gene Number and Gene Density 38 Very Different Gene Densities in E. coli, Yeast, Drosophila and Human 39 Organism Complexity and Gene Numbers Mycoplasma 1 Cell (Bacterium) E. Coli Yeast C. elegans Drosophila 1 Cell (Bacterium) 1 Cell 1,000 Cells 10,000 Cells 470 Genes 4290 Genes 6241 Genes 19,000 Genes 14,000 Genes 24,500 Genes 23,000 Genes Arabidopsis 50 Cell types Human Trillions of Cells 40 Features of Human Proteome 41 Comparative Proteomics 42 Comparative Proteomics Genome-wide comparison of transcriptional activator families in eukaryotes 43 Topic 4 Microarray 44 Genome, Transcriptome and Proteome Proteomics Genomics Metabolomics (Chemogenomics ) DNA RNA PROTEIN METABOLITE Genome Transcriptome Proteome 45 Making a Microarray by Microspotting 46 Preparing a DNA Microarray by Microspotting 47 Detection of Gene Expression by Microarray 48 Results of a Real Microarray Experiment 49 Preparing a DNA Microarray by Photolithography 50 Gene Chip 51 Using a DNA Microarray 52 Global Gene Expression Analysis Using DNA Microarray 53 Animation 2: Microarray 54 Summary Genomics is the science of mapping, sequencing, comparison and functional analysis of the entire genomes of organisms. Map-based genome sequencing uses orderly mapped DNA fragments for sequencing. The assembly of sequencing data is straightforward. Shotgun genome sequencing uses random, overlapping DNA fragments for sequencing. The assembly of sequencing data needs special algorithm and enormous computing power. DNA microarray is a new technique for gathering large amount of genetic data in parallel. 55 ...
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