Lecture #9Date _____Chapter 20~DNA Technology & Genomics

O.J. Simpson capital murder case,1/95-9/95

Odds of blood in Ford Bronco not being R. Goldmans:6.5 billion to 1Odds of blood on socks in bedroom not being N. Brown-Simpsons:8.5 billion to 1Odds of blood on glove not being from R. Goldman, N. Brown-Simpson, and O.J. Simpson:21.5 billion to 1Number of people on planet earth:6.1 billionOdds of being struck by lightning in the U.S.:2.8 million to 1Odds of winning the Illinois Big Game lottery:76 million to 1 Odds of getting killed driving to the gas station to buy a lottery ticket 4.5 million to 1Odds of seeing 3 albino deer at the same time:85 million to 1Odds of having quintuplets:85 million to 1Odds of being struck by a meteorite:10 trillion to 1

Recombinant DNADNA in which genes from 2 different sources are linkedGenetic engineering: direct manipulation of genes for practical purposesBiotechnology: manipulation of organisms or their components to perform practical tasks or provide useful products

Bacterial plasmids in gene cloning

DNA CloningRestriction enzymes (endonucleases):in nature, these enzymes protect bacteria from intruding DNA; they cut up the DNA (restriction); very specificRestriction site:recognition sequence for a particular restriction enzymeRestriction fragments:segments of DNA cut by restriction enzymes in a reproducable waySticky end:short extensions of restriction fragmentsDNA ligase: enzyme that can join the sticky ends of DNA fragmentsCloning vector: DNA molecule that can carry foreign DNA into a cell and replicate there (usually bacterial plasmids)

Steps for eukaryotic gene cloningIsolation of cloning vector (bacterial plasmid) & gene-source DNA (gene of interest)Insertion of gene-source DNA into the cloning vector using the same restriction enzyme; bind the fragmented DNA with DNA ligaseIntroduction of cloning vector into cells (transformation by bacterial cells)Cloning of cells (and foreign genes)Identification of cell clones carrying the gene of interest

DNA Analysis & GenomicsPCR (polymerase chain reaction)Gel electrophoresisRestriction fragment analysis (RFLPs)Southern blottingDNA sequencingHuman genome project

Polymerase chain reaction (PCR)Amplification of any piece of DNA without cells (in vitro)Materials: heat, DNA polymerase, nucleotides, single-stranded DNA primersApplications: fossils, forensics, prenatal diagnosis, etc.

DNA AnalysisGel electrophoresis: separates nucleic acids or proteins on the basis of size or electrical charge creating DNA bands of the same length

Restriction fragment analysisRestriction fragment length polymorphisms (RFLPs)Southern blotting: process that reveals sequences and the RFLPs in a DNA sequenceDNA Fingerprinting

DNA SequencingDetermination of nucleotide sequences (Sanger method, sequencing machine)Genomics: the study of genomes based onDNA sequencesHuman Genome Project

Practical DNA Technology UsesDiagnosis of diseaseHuman gene therapyPharmaceutical products (vaccines)ForensicsAnimal husbandry (transgenic organisms)Genetic engineering in plantsEthical concerns?

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