Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Chapter 11 Lecture Outline See PowerPoint Image Slides for all figures and tables pre-inserted into PowerPoint without notes.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Why Biotechnology Works Biotechnology is a collection of laboratory techniques that involve the direct manipulation of an organism’s DNA. Biotechnology has allowed for: – Cheaper and more effective drugs – The correction of genetic mutation – The creation of cells that can clean-up environmental messes – An increase in agricultural productivity

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Understanding Biotechnology Biotechnology is based on our understanding of the structure of DNA. In the nucleus, chromosomes are made of – DNA – Histone proteins DNA is the genetic material of the cell. – The information is in the sequence of nucleotides in the DNA. – Genes are regions of DNA that encode for specific proteins. Proteins produce the characteristics of cells (phenotype). Biotechnology involves the manipulation of DNA in order to change phenotype.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Comparing DNA Organisms that have similar phenotypes have similar DNA sequences. DNA between two organisms can be compared in a couple of different ways: – DNA fingerprinting – DNA sequencing

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Fingerprinting Uniquely identifies individuals on the basis of DNA fragment lengths. – Fragments are generated by restriction enzymes that cut DNA at specific sites. – Each individual’s DNA is different enough that these enzymes will generate different lengths of fragments in two different individuals.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Variable Number Tandem Repeats (VNTR) Variable number tandem repeats are regions of DNA that are repetitive sequences. Each person has a slightly different number of repeats. Therefore, if these regions are cut with restriction enzymes, each person will have a different set of fragments.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Other Techniques Used in DNA Fingerprinting Polymerase chain reaction – DNA replication in a test tube – Uses primers that dictate the region of DNA to be copied – Makes many copies of a particular segment of DNA – This creates enough material for scientists to perform DNA fingerprinting.

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Other Techniques Used in DNA Fingerprinting Electrophoresis – A way to separate DNA fragments based on their length – DNA sample is loaded into a gel matrix and an electrical current is applied. – Smaller fragments travel through the gel faster. – Creates a banding pattern of fragments

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Electrophoresis

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Fingerprinting Technique DNA is obtained from two sources: – Blood at a crime scene – Hair from the crime suspect Polymerase chain reaction is used to make many copies of a VNTR region. Restriction enzymes are used to cut the VNTRs into fragments. The fragments are separated by electrophoresis. The patterns of the two DNA samples are compared.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Fingerprinting Applications DNA fingerprinting can identify if two samples of DNA came from the same person. Used in the prosecution of crime suspects Used in paternity cases

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Fingerprinting in a Crime Case

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Fingerprinting in a Paternity Case

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display DNA Sequencing A specially engineered DNA replication reaction. – The DNA of interest is the template. – Uses DNA polymerase – Uses a primer (gives DNA polymerase a place to start) – Uses regular nucleotides (A,T,G,C) – Uses special fluorescently-labeled dideoxynucleotides When these are added to the growing chain, replication will stop. The fragment will be fluorescently labeled as well. – Fragments are separated by electrophoresis, and the sequence can be read.

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display The Human Genome Project A massive collaborative effort to determine the sequence of DNA in all human chromosomes. Started with physical maps of chromosomes – Determines the location of specific markers on the chromosomes DNA sequencing determined the exact nucleotide sequence between each marker.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Genes Known to be on Human Chromosome 21

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Human Genome Project Applications Disease diagnosis Discovery of new family of proteins Better understanding of basic biology The human genome project found that there are far fewer genes in the human genome than previously predicted. – Each gene can be alternatively spliced to code for several proteins.

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Other Genomes The genomes of other organisms have also been sequenced.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Patterns in Protein Coding The human genome was compared to these other genomes. – Several types of patterns were found. Tandem clusters  grouped copies of the same gene Segmental duplications  groups of genes copied from one chromosome and moved to another chromosome Multigene families are groups of different genes that are closely related. – The parts of these proteins that are similar represent important regions of the proteins that are crucial to their function.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Patterns in Protein Coding Sequences

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display New Fields of Knowledge Recent developments in genome sequencing have led to three new fields in biology. – Genomics The comparison of genomes from different species Identifies species relatedness and gene similarities – Transcriptomics Examines when, where and how much mRNA for a given gene is made – Proteomics Examines the proteins that are predicted from the DNA sequence

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Genetic Modification of Organisms It is now possible to clone genes and move them from one organism to another. – Called gene cloning DNA sequences can be altered (mutated) to generate a desired change. – The new DNA is called recombinant DNA. Once the DNA is transferred, the new host cell begins to make the new DNA and produce the new proteins. – Organisms that contain recombinant DNA are called “genetically modified organisms”. – Usually involves bacteria or viruses that will make large amounts of the protein of interest

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display How to Clone a Gene Cut the gene of interest out of the chromosome using restriction enzymes.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display How to Clone a Gene Splice the fragment containing the gene into a carrier molecule, usually a bacterial plasmid.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display How to Clone a Gene Insert the plasmid with the fragment into the bacterial cells.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display How to Clone a Gene Each time the bacterial cells divide, many copies of the gene will be made.

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Genetically Modified Organisms Genetically modified organisms have been used to: – Make human insulin – Generate “Golden rice” – Make interferon – Make human growth hormone

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Genetically Modified Organisms – For bioremediation (the use of living organisms to remove toxins from the environment) – Generate crops that supply developing nations with nutrients not normally found in their native plants – Generate crops that can manufacture medicines to treat disease – Generate crops that are resistant to herbicides or that make their own insecticides

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Gene Therapy Recombinant DNA technology can be used to administer gene therapy. – Gene therapy involves manipulating genes in order to cure or treat a genetic disease.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Gene Therapy Gene therapies must be specifically designed for each situation. – If the mutant gene is not functional, then a functional gene must be inserted. – If the mutant gene is overactive, then it must be deleted or altered. Usually involves mutating the part of the gene that controls its activation

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display The Process of Gene Therapy

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Cloning Organisms Recombinant DNA technologies have also enabled scientists to clone entire organisms. Cloning in the laboratory involves a process called somatic cell nuclear transfer. – First, a nucleus must be removed from a cell of the organism that is to be cloned. – The nucleus is then placed into an enucleated egg. – The egg is stimulated to divide and is artificially implanted into a host mother. Several species have been successfully cloned. – Sheep, cats, monkeys, etc.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Cloning Dolly, the Sheep

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display The Scientific Relevance of Cloning Cloning has advanced our understanding of developmental processes. – Determination is the process a cell goes through to select which genes it will express. – Differentiation is the process of expressing those genes and becoming a specific cell type. The cloning of Dolly illustrated that determination can be reversed and a fully differentiated cell can be used to clone an organism.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Stem Cells Stem cells have not completed the determination and differentiation process, so they have the potential to develop into many different cell types. – Stem cells could be used to replace damaged tissues and organs in humans. – Could be used to cure diseases Parkinson’s disease Diabetes

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Embryonic Stem Cells Embryonic stem cells come from embryos and have not completed the process of determination. – These have great potential. – However, embryos must be destroyed in order to obtain them and work with them.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display The Culturing of Embryonic Stem Cells

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Adult Stem Cells Adult stem cells have partially completed the process of determination. – These have less potential; they are already specific tissue types (blood, bone, etc). – There is no ethical dilemma in obtaining them.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display The Differentiation of Blood Cells

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Personalized Stem Cell Lines Combining somatic cell nuclear transfer and stem cells may allow scientists to create personalized stem cell lines. – The individual’s nucleus would be transferred to an egg. – The egg would be allowed to divide to the point where it produced embryonic stem cells. – Could potentially be used to grow tissues or organs (in the future; not possible now).

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Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Biotechnology Ethics The recent advances in biotechnology have raised a number of ethical questions. – Will the technology be used safely? – Who will benefit? – Who will suffer? – Should the technology be used to make a profit? – Just because we can, should we?

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display What are the consequences? One approach to ethics is to weigh the “pros” and “cons” of the particular technology or application. – Genetic testing allows us to predict disease before it happens. Pro: This allows one to seek treatment early. Con: If insurance companies obtain this information, they may deny coverage.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display What are the consequences? – Cloning technology allows us to generate nutritionally advanced foods. Pro: This provides underdeveloped nations with better food sources. Con: The GMO organisms may impact the ecosystem negatively.

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Is it inherently wrong? Another approach to ethics is to ask if the technology and its application violates principles that are valued by society. – Does it threaten someone’s human rights? – Does it threaten someone’s Bill of Rights? – Does it violate someone’s religious beliefs? – Does it diminish someone’s quality of life? – Does it violate animal rights?

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display Is it inherently wrong? Is it inherently wrong to… – Produce genetically modified organisms? – Manipulate genes? – Manipulate or destroy embryos to obtain stem cells?