Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Lectures by Chris C. Romero PowerPoint ® Lectures for Essential Biology, Third Edition – Neil Campbell, Jane Reece, and Eric Simon Essential Biology with Physiology, Second Edition – Neil Campbell, Jane Reece, and Eric Simon CHAPTER 12 DNA Technology

Biology and Society: Crime Scene Investigations: Murders in a Small Town On November 22, 1983, –A 15-year-old girl was raped and murdered on a quiet country lane. –Three years later, another 15-year-old girl was raped and murdered. Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

DNA fingerprinting of DNA samples from suspects and the crime scene –Proved one man guilty and another man innocent.

Figure 12.1

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant DNA Technology Recombinant DNA technology is a set of techniques for combining genes from different sources into a single DNA molecule. –An organism that carries recombinant DNA is called a genetically modified (GM) organism. Recombinant DNA technology is applied in the field of biotechnology. –Biotechnology uses various organisms to perform practical tasks.

Figure 12.2

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings From Humulin to Genetically Modified Foods By transferring the gene for a desired protein product into a bacterium, proteins can be produced in large quantities.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Making Humulin In 1982, the world’s first genetically engineered pharmaceutical product was produced. –Humulin, human insulin, was produced by genetically modified bacteria.

Unnumbered Figure p. 222

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Humulin was the first recombinant DNA drug approved by the FDA.

Figure 12.3

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings DNA technology is also helping medical researchers develop vaccines. –A vaccine is a harmless variant or derivative of a pathogen. –Vaccines are used to prevent infectious diseases.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Genetically Modified (GM) Foods Today, DNA technology is quickly replacing traditional plant-breeding programs. –In the United States today, roughly one-half of the corn crop and over three-quarters of the soybean and cotton crops are genetically modified.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Corn has been genetically modified to resist insect infestation. –This corn has been damaged by the European corn borer.

Figure 12.4

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings “Golden rice” has been genetically modified to contain beta-carotene. –Our bodies use beta-carotene to make vitamin A.

Figure 12.5

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Farm Animals and “Pharm” Animals While transgenic plants are used today as commercial products, transgenic whole animals are currently only in the testing phase. These transgenic sheep carry a gene for a human blood protein. –This protein may help in the treatment of cystic fibrosis.

Figure 12.6

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings While transgenic animals are currently used to produce potentially useful proteins, none are yet found in our food supply. It is possible that DNA technology will eventually replace traditional animal breeding.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant DNA Techniques Bacteria are the workhorses of modern biotechnology. To work with genes in the laboratory, biologists often use bacterial plasmids. –Plasmids are small, circular DNA molecules that are separate from the much larger bacterial chromosome.

Figure 12.7

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Plasmids can easily incorporate foreign DNA. Plasmids are readily taken up by bacterial cells. –Plasmids then act as vectors, DNA carriers that move genes from one cell to another.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant DNA techniques can help biologists produce large quantities of a desired protein.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Glow in the Dark Fish

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Review 1.What is Recombinant DNA technology? 2.What is needed (ingredients)? 3.What are applications of this process? What you need to Know about DNA Technology. 1. Technique 2. Purpose 3. Pros/Cons

Figure 12.8

A Closer Look: Cutting and Pasting DNA with Restriction Enzymes Recombinant DNA is produced by combining two ingredients: –A bacterial plasmid –The gene of interest To combine these ingredients, a piece of DNA must be spliced into a plasmid. Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

This splicing process can be accomplished using restriction enzymes. –These enzymes cut DNA at specific nucleotide sequences. These cuts produce pieces of DNA called restriction fragments –That may have “sticky ends” that are important for joining DNA from different sources.

Figure 12.9

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Read background information and underline answers to: -What is recombinant DNA? -What is a plasmid? -What occurs at the origin of replication? -What are ampicillin and kanamycin? -What are pAMP and pKAN? -What role do bacteria play? Read background information and underline answers to: -What is recombinant DNA? -What is a plasmid? -What occurs at the origin of replication? -What are ampicillin and kanamycin? -What are pAMP and pKAN? -What role do bacteria play? /content/plasmidcloning.html

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Bacterial growth without plasmid: No antibiotics AmpicillinKanamycin Ampicillin& Kanamycin

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Bacterial growth without plasmid: No antibiotics AmpicillinKanamycin Ampicillin& Kanamycin

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Bacterial growth with plasmid: No antibiotics AmpicillinKanamycin Ampicillin& Kanamycin

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Bacterial growth with plasmid: No antibiotics AmpicillinKanamycin Ampicillin& Kanamycin

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant Paper Plasmid lab 1.pAMP = 1A (amp) and 1B pKAN = 2 A (kan) and 2B 1A + 1B (resistant to amp) 1B + 2B (no resistance) 1A + 2B (resistant to amp) 2A + 1B (no origin of rep) 1A + 2A (resistant to both) 2A + 2B (resistant to kan)

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant paper plasmid lab 2. No antibiotics + plasmid Amp + plasmid Kan + plasmid Amp/Kan + plasmid No antibiotics - plasmid Amp - plasmid Kan - plasmid Amp/Kan - plasmid

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant Paper Plasmid lab 3. pAMP = 1A (amp) and 1B pKAN = 2 A (kan) and 2B Would survive on one antibiotic but not the other! 1A + 1B (resistant to amp) 1A + 2B (resistant to amp) 2A + 2B (resistant to kan) 2A + 1B (resistant to kan, but no origin of rep)

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Recombinant Paper Plasmid Lab 4.The most important reason is so that scientists can identify bacteria that have been transformed. - Can select only bacteria that have the gene of interest!

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings A Closer Look: Obtaining the Gene of Interest How can a researcher obtain DNA that encodes a particular gene of interest? The “shotgun” approach is one way to synthesize a gene of interest. –Millions of recombinant plasmids containing different segments of foreign DNA are produced. –This collection is called a genomic library.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Once a genomic library is created, biologists must identify the bacterial clone containing the desired gene. –A specific sequence of radioactive nucleotides matching those in the desired gene can be created. –This type of labeled nucleic acid molecule is called a nucleic acid probe.

Figure 12.10

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Reverse transcriptase is another method for synthesizing a gene of interest.

Figure 12.11

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings DNA Profiling and Forensic Science DNA technology has rapidly revolutionized the field of forensics. –Forensics is the scientific analysis of evidence from crime scenes. DNA profiling can be used to determine whether or not two samples of genetic material are from a particular individual.

Figure 12.12

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Murder, Paternity, and Ancient DNA DNA profiling (fingerprinting) –Has become a standard criminology tool. –Has been used to identify victims of the September 11, 2001, World Trade Center attack. –Can be used in paternity cases.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings DNA fingerprinting is also used in evolutionary research –To study ancient pieces of DNA, such as that of Cheddar Man. –Found in a cave near Cheddar, England –Direct ancestor of local teacher

Figure 12.13

DNA Fingerprinting Techniques The Polymerase Chain Reaction (PCR) The polymerase chain reaction (PCR) is a technique by which any segment of DNA can be copied quickly and precisely. –Through PCR, scientists can obtain enough DNA from even minute amounts of blood or other tissue to allow DNA profiling. Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Figure 12.14

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Short Tandem Repeat (STR) Analysis How do you prove that two samples of DNA come from the same person?

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Short tandem repeats (STRs) –Are repetitive sequences of DNA that are repeated various times in the genome.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Scientists use STR analysis –To compare the number of repeats between different samples of DNA.

Figure 12.15

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Once a set of DNA fragments is prepared, –The next step in STR analysis is to determine the lengths of these fragments.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Gel Electrophoresis –Can be used to separate the DNA fragments obtained from different sources.

Figure 12.16

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings The DNA fragments are visualized as “bands” on the gel. –The bands of different DNA samples can then be compared.

Figure 12.17

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings One common application of gel electrophoresis is RFLP analysis, –In which DNA molecules to be compared are exposed to a series of restriction enzymes.

Figure 12.18

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Genomics and Proteomics Genomics is the science of studying whole genomes. –The first targets of genomics were bacteria.

Table 12.1

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings The Human Genome Project In 1990, an international consortium of government-funded researchers began the Human Genome Project. –The goal of the project was to sequence the human genome.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Sequencing of the human genome presented a major challenge. –It is very large. –Only a small amount of our total DNA is contained in genes that code for proteins.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings The Human Genome Project –Can help map specific disease genes such as Parkinson’s disease.

Figure 12.20

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Genome-Mapping Techniques The Human Genome Project proceeded through several stages, –During which preliminary maps were created and refined.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings The whole-genome shotgun method –Involves sequencing DNA fragments from an entire genome and reassembling them in a single stage.

Figure 12.21

The Process of Science: Can Genomics Cure Cancer? In 2003, the Food and Drug Administration –Approved the drug gefinitib for the treatment of lung cancer. Unfortunately, gefinitib is ineffective for many patients. Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings

A 2004 study found that genetic differences among patients affected their response to the drug, –Exhibiting how genomics can affect the treatment of disease.

Figure 12.22

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Proteomics Success in genomics has given rise to proteomics, –The systematic study of the full set of proteins found in organisms.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Human Gene Therapy Human gene therapy is a recombinant DNA procedure that seeks to treat disease by altering the genes of the afflicted person. –The mutant version of a gene is replaced or supplemented with a properly functioning one.

Figure 12.23

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Treating Severe Combined Immunodeficiency SCID is a fatal inherited disease caused by a single defective gene. –The gene prevents the development of the immune system. –SCID patients quickly die unless treated with a bone marrow transplant.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Since the year 2000, –Gene therapy has successfully cured 22 children with inborn SCID. Unfortunately, three of the children developed leukemia and one of them died.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Safety and Ethical Issues As soon as scientists realized the power of DNA technology, they began to worry about potential dangers such as: –The creation of hazardous new pathogens –The transfer of cancer genes into infectious bacteria and viruses

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Strict laboratory safety procedures have been designed to protect researchers from infection by engineered microbes. –Procedures have also been designed to prevent microbes from accidentally leaving the laboratory.

Figure 12.24

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings The Controversy over Genetically Modified Foods GM strains account for a significant percentage of several agricultural crops in the United States.

Figure 12.25

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Advocates of a cautious approach have some concerns: –Crops carrying genes from other species might harm the environment. –GM foods could be hazardous to human health. –Transgenic plants might pass their genes to close relatives in nearby wild areas.

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Negotiators from 130 countries (including the United States) agreed on a Biosafety Protocol. –The protocol requires exporters to identify GM organisms present in bulk food shipments. Several U.S. regulatory agencies evaluate biotechnology projects for potential risks: –Department of Agriculture –Food and Drug Administration –Environmental Protection Agency –National Institutes of Health

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Ethical Questions Raised by DNA Technology Should genetically engineered human growth hormone be used to stimulate growth in HGH- deficient children?

Figure 12.26

Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings Genetic engineering of gametes and zygotes has been accomplished in lab animals. –Should we try to eliminate genetic defects in our children? –Should we interfere with evolution in this way? Advances in genetic fingerprinting raise privacy issues. What about the information obtained in the Human Genome Project? –How do we prevent genetic information from being used in a discriminatory manner?

Evolution Connection: Genomes Hold Clues to Evolution Genome data has confirmed evolutionary connections. Comparisons of completed genome sequences strongly support the theory that there are three fundamental domains of life: –Bacteria –Archaea –Eukaryotes Copyright © 2007 Pearson Education Inc., publishing as Pearson Benjamin Cummings