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Chapter 9 Genetic engineering • Deliberate manipulation of genes in an organism. • Done in a lab by scientists • Therapeutic substances such as human insulin. • Insulin came from pancreas obtained from slaughtered animals • Genetically engineered E.coli cells to make human insulin Tools used in genetic engineering Restriction enzymes • Bacteria – restriction enzymes • To breakdown phage DNA • Extract the restriction enzyme from bacteria and use it genetic engineering • EcoRI, BamHI – recognize specific sequence • Staggered cuts on DNA • Ends of the fragment are single stranded Sticky ends Cohesive ends ligase • Vectors – carry the gene of interest into a bacterial cell. • Plasmids • Small enough – they can enter into the cell • Selection markers – antibiotic resistance genes. • Help in selecting the cells that have the gene of interest. Gene library • • • • • Collection of genes from an organism Mouse gene library, yeast gene library Biotechnology company Yeast gene library DNA – fragment – restriction enzyme – fragments are inserted into plasmids – introduced into a bacterial cell • Each bacterial cell having the recombinant cell is a clone • Large # clones – a clone for each gene that exists in the yeast cell Hexokinase Malate Dehydrogebase pheromone Helps the cells To come together And mate cDNA • • • • • • Complementary DNA Eukaryotic genes – introns and exons Introns – noncoding regions Exons - coding regions cDNA – synthetic gene – has only exons c stands for complementary DNA • If we want to introduce eukaryotic gene into a prokaryotic cell, we should use cDNA • If we place natural eukaryotic gene into a • Bacterial cell, it cannot remove the introns. • Functional protein will not be produced by the prokaryotic cell. Applications of genetic engineering • Hormones – insulin • Growth hormone – somatotorpin • Produced by genetically engineered E.coli cells. • Treat stunted growth • Pituitary gland removed during autopsy. vaccine • Hepatitis B vaccine • Genetically engineered Saccharomyces cells • Vaccine has only the protein part of the virus. • Does not have the genetic material of the virus • No chance of getting the disease due to vaccination Genetic screening • Carrier of genetic disorder • Cystic fibrosis genetic disorder • Mutation in a gene that codes for a membrane protein. • Protein is not functional • Produce thick mucus blocks airways and causes various health problems. • Person has the disease – symptoms • Some people are carriers • One cystic fibrosis gene (father) one normal gene (mother) – heterozygous for cystic fibrosis • Two carriers – have a baby – the baby can have the disease • Genetic screening is used – procedure known as southern blotting. Gel electrophoresis Single stranded DNA complementary hybridize Agriculture Forensic medicine Each person has a unique set of introns. restriction enzyme fragment pattern of DNA Polymerase Chain Reaction Amplify DNA Target DNA Nucleotides DNA polymerase