lecture 27 W11 - Lecture 27 (3/21/11) - Genomics Today's...

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Today’s topics 1) Revolution in large scale DNA sequencing 2) What has the sequence of genomes told us. 3) How did gene families arise. 4) The use of microarrays to measure the transcriptome of a cell. Lecture 27 (3/21/11) - Genomics
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Molecules; Chemical bonds; Free energy Lipids Amino acids nucleotides carbohydrates membranes proteins Membrane transport Membrane proteins cytoskeleton enzymes DNA RNA Metabolism Sugar transport Electrolyte transport Kidney Nerve cells Transporters and pumps Receptors Cell-cell communication Chemical signaling development Replication transcription translation Glycolysis Fermentation Krebs cycle Photo- Synthesis & Calvin cycle Cell cycle Regulation of transcription biotechnology genomics Microbes Biology 172 flowchart (lecture 27) cancer epigenetics
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SHOTGUN SEQUENCING A GENOME 160 kb fragments Genomic DNA BAC library 1-kb fragments “Shotgun clones” BAC Main bacterial chromosome 1. Cut DNA into fragments of 160 kb, using sonication. 2. Insert fragments into bacterial artificial chromosomes; grow in E. coli cells to obtain large numbers of each fragment. 3. Purify each 160-kb fragment, then cut each into a set of 1-kb fragments, using sonication, so that 1-kb fragments overlap. 4. Insert 1-kb fragments into plasmids; grow in E. coli cells. Obtain many copies of each fragment. Shotgun sequences 5. Sequence each fragment. Find regions where different fragments overlap. Draft sequence 6. Assemble all the 1-kb fragments from each original 160-kb fragment by matching overlapping ends. 7. Assemble sequences from different BACs (160-kb fragments) by matching overlapping ends. SHOTGUN SEQUENCING A GENOME
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Shotgun sequences 5. Sequence each fragment. Find regions where different fragments overlap. Draft sequence 6. Assemble all the 1-kb fragments from each original 160-kb fragment by matching overlapping ends. 7. Assemble sequences from different BACs (160-kb fragments) by matching overlapping ends. SHOTGUN SEQUENCING A GENOME Finished product (contig)
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Computer programs are used to search a genome sequence in both directions and identify open reading frames (ORFs) . ORFs are identified based on long stretches of sequence that lack a stop codon but are flanked by a start codon and a stop codon.
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There are no introns in bacteria and archaea, and genes can be identified by highly conserved promoter sequences associated with a distinct ORF. In eukaryotic organisms, genes contain introns, and identification of ORFs is more difficult. One strategy is to use cDNA libraries to identify mRNA sequences and then locate the gene in the genome sequence.
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Why do you think that it is necessary to use multiple cDNA libraries are necessary to aid in the identification of genes
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This note was uploaded on 02/13/2012 for the course BIO 172 taught by Professor Clark during the Winter '08 term at University of Michigan.

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lecture 27 W11 - Lecture 27 (3/21/11) - Genomics Today's...

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