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Genomics Study of the Functions and Interactions of Many Genes at Once Goal of the Human Genome Project: To Determine the Sequence of All of the DNA in a Human Cell Per haploid genome Now estimated at 30,000 genes How Can We Use Human Genome Information? Cytogenetic map or Distances: 1 base Distances: 10,000 bases Distances: 100,000 bases Distances: 5,000,000 bases Chromosomal Maps From A B A B D C B B E To B D D E E E C F F Distance in cM 20 8 12 6 14 9 15 28 22 Determining the Human Genome Sequence Sequencing: The order of DNA nucleotides is determined for fragments produced by restriction enzymes Assembly: Computer analysis determines the order of fragments by aligning overlapping regions Assembly: Using Overlapping Regions to Arrange Sequences Determining the Human Genome Sequence Annotation: Computer analysis locates possible gene regions and identifies potential functions Gene Expression: Studying RNA production by Microarray Analysis to identify protein-coding sequences in specific cell types DNA Sequencing by the Sanger Method Relies on Chain Termination during replication due to incorporation of Dideoxynucleotides OH H H H deoxyribose dideoxyribose Replicate unknown DNA using dideoxynucleotides to terminate growing chains at various positions Unknown Sequence 3’-A-G-C-C-C-A-G-A-T-T-5’ Complementary Products Black= dideoxynucleotide 5’-T-3’ 5’-T-C-G-G-G-3’ 5’-T-C-G-3’ 5’-T-C-G-G-G-T-C-T-3’ 5’-T-C-3’ 5’-T-C-G-G-G-T-C-T-A-3’ 5’-T-C-G-G-G-T-C-T-A-A-3’ 5’-T-C-G-G-G-T-C-3’ 5’-T-C-G-G-G-T-3’ 5’-T-C-G-G-3’ Use gel electrophoresis to separate replicated fragments that differ by one base A Dideoxynucleotide T C G Size of DNA on gel shows distance from start of replication Labeling of dideoxynucleotide shows complementary sequence Determine unknown sequence from complementary sequence with base-pairing rules A Dideoxynucleotide T C G Complementary Sequence 5’-T-C-G-G-G-T-C-T-A-A 5’-T-C-G-G-G-T-C-T-A 5’-T-C-G-G-G-T-C-T 5’-T-C-G-G-G-T-C 5’-T-C-G-G-G-T 5’-T-C-G-G-G 5’-T-C-G-G 5’-T-C-G 5’-T-C 5’-T Complementary Sequence Unknown Sequence 5’-T-C-G-G-G-T-C-T-A-A-3’ 3’-A-G-C-C-C-A-G-A-T-T-5’ Use of Fluorescent Dideoxynucleotides in Sanger Sequencing Microarray Analysis • Microarray • DNA sequences arranged in a matrix • Tests for binding to complementary sequences • Steps in Microarray Analysis • Arrange DNA or cDNA in an ordered array on a glass slide • Incubate array with fluorescently-labeled probe • Detect probe hybridization using a laser • Record and store data with computers Array of 1046 cDNAs probed with bone marrow based cDNA probe Using Human Genome Information Pharmacogenomics Using DNA Microarrays to select individual drug treatments Mutations in Gene for Affect the following disease conditions Glutathione-STransferase (GST) Detoxification of environmental toxins Susceptibility to some cancers Cytochrome P450 Affects rate of metabolizing 30-40 drugs 5-10% of Caucasians and African Americans have low rates Apo E4 Gene dose affects Age of onset for Alzheimer's disease Response to drug Tacrine