The following will be discussed in this chapter: What are the methods that scientists use to create recombinant DNA molecules; can scientists create genes from recombinant DNA molecules; and can scientists modify the heredity of an organism using recombinant DNA? | Chapter 13 Recombinant DNA: Cloning and Creation of Chimeric Genes to accompany Biochemistry, 2/e by Reginald Garrett and Charles Grisham All rights reserved. Requests for permission to make copies of any part of the work should be mailed to: Permissions Department, Harcourt Brace & Company, 6277 Sea Harbor Drive, Orlando, Florida 32887-6777 Outline Cloning DNA Libraries Polymerase Chain Reaction (PCR) Recombinant DNA Technology Cloning Clone: a collection of molecules or cells, all identical to an original molecule or cell To "clone a gene" is to make many copies of it - for example, in a population of bacteria Gene can be an exact copy of a natural gene Gene can be an altered version of a natural gene Recombinant DNA technology makes it possible Plasmids Naturally occurring extrachromosomal DNA Plasmids are circular dsDNA Plasmids can be cleaved by restriction enzymes, leaving sticky ends Artificial plasmids can be constructed by linking new DNA fragments to the sticky ends of plasmid Cloning Vectors Plasmids that can be modified to carry new genes Plasmids useful as cloning vectors must have a replicator (origin of replication) a selectable marker (antibiotic resistance gene) a cloning site (site where insertion of foreign DNA will not disrupt replication or inactivate essential markers Chimeric Plasmids Named for mythological beasts with body parts from several creatures After cleavage of a plasmid with a restriction enzyme, a foreign DNA fragment can be inserted Ends of the plasmid/fragment are closed to form a "recombinant plasmid" Plasmid can replicate when placed in a suitable bacterial host See Figure Directional Cloning Often one desires to insert foreign DNA in a particular orientation This can be done by making two cleavages with two different restriction enzymes Construct foreign DNA with same two restriction enzymes Foreign DNA can only be inserted in one direction See Figure DNA Libraries Sets of cloned DNA . | Chapter 13 Recombinant DNA: Cloning and Creation of Chimeric Genes to accompany Biochemistry, 2/e by Reginald Garrett and Charles Grisham All rights reserved. Requests for permission to make copies of any part of the work should be mailed to: Permissions Department, Harcourt Brace & Company, 6277 Sea Harbor Drive, Orlando, Florida 32887-6777 Outline Cloning DNA Libraries Polymerase Chain Reaction (PCR) Recombinant DNA Technology Cloning Clone: a collection of molecules or cells, all identical to an original molecule or cell To "clone a gene" is to make many copies of it - for example, in a population of bacteria Gene can be an exact copy of a natural gene Gene can be an altered version of a natural gene Recombinant DNA technology makes it possible Plasmids Naturally occurring extrachromosomal DNA Plasmids are circular dsDNA Plasmids can be cleaved by restriction enzymes, leaving sticky ends Artificial plasmids can be constructed by linking new DNA fragments to
