Concepts and Applications Seventh Edition Powerpoint Lecture Outline Human Genetics Concepts and Applications Seventh Edition Ricki Lewis Prepared by Mary King Kananen Penn State Altoona

Chapter 19 Genetically Modified Organisms

Transgenic Animals Have genetic modifications and carry that genetic alteration from other organisms in all of their cells Example Genetically modified pigs were engineered to produce less polluting manure Chapter Opener 19

Biotechnology Use or alteration of cells or biological molecules for specific application Transgenic organisms are possible, but the genetic code is universal Ethical and legal issues to be considered including patent laws Figure 19.1

Technology Timeline

Amplifying DNA Polymerase chain reaction (PCR) Molecular Increases the amount of a DNA sequence Replicates sequence millions of times Recombinant DNA technology Amplifies DNA that includes Within cells Sequences from other organisms

Uses of PCR Table 19.1

Figure 19.2

Transcription-Mediated Amplification Copies target DNA into RNA and then uses RNA polymerase to amplify RNA Does not require temperature shifts Makes 10 billion copies in ½ hour Used to develop a test for HIV, sensitive enough to detect early infections

Recombinant DNA Recombinant DNA is a molecule that combines DNA from two sources Also known as gene cloning Creates a new combination of genetic material Human gene for insulin was placed in bacteria The bacteria are recombinant organisms and produce insulin in large quantities for diabetics Genetically modified organisms are possible because of the universal nature of the genetic code

Creating Recombinant DNA Molecules Cut DNA from donor and recipient with the same restriction enzymes Cut DNA fragment is combined with a vector Vector DNA moves and copies DNA fragment of interest Vector cut with restriction enzymes The complementary ends of the DNAs bind and ligase enzyme reattaches the sugar-phosphate backbone of the DNA

Creating Recombinant DNA Molecules Figure 19.3a

Figure 19.3b

Figure 19.3c

Vectors Are DNA molecules that can be moved into and replicated in an organism They are classified by The organisms that replicated the vector The size of DNA that can be inserted Table 19.2

Plasmids Figure 19.4

Recombinant DNA Figure 19.5

Isolating Gene of Interest Genomic library Collections of recombinant DNA that contain pieces of the genome DNA probe Radioactively labeled gene fragments cDNA library Genomic library of protein encoding genes produced by extracting mRNA

Figure 19.6

Selecting Recombinant Molecules Three types of cells can result from attempt to introduce a DNA molecule into a bacterial cell: Cells lack plasmid Cells contain plasmid that do not contain foreign genes Cells that contain plasmids with foreign genes

Selecting for Cells with Vectors Vectors are commonly engineered to carry antibiotic resistance genes Host bacteria die in the presence of the antibiotic Bacteria harboring the vector survive Growing cells on media with antibiotics ensures that all growing cells must carry the vector

Figure 19.7

Figure 19.8

Selecting Cells with Inserted DNA The site of insertion of the DNA of interest can be within a gene on the vector Insertion of a DNA fragment will disrupt the vector gene The gene lacZ produces an enzyme and bacteria to turn blue in the presence of certain media Insertions in the lacZ gene prevent lacZ enzyme production and the bacteria are white Bacteria with vectors that are white carry an DNA inserted in the lacZ gene Rare mutations in the lacZ gene will also make white bacteria

Applications of Recombinant DNA Recombinant DNA is used to: Study the biochemical properties or genetic pathways of that protein Mass produce a particular protein (e.g., insulin) Sometimes conventional methods are still the better choice Textile industry can produce the dye indigo in E. coli by genetically modifying genes of the glucose pathway and introducing genes from another bacterial species

Table 19.3

The Dye Indigo The blue in blue jeans Originally came from mollusks or fermented leaves of woad or indigo plants Synthetic process uses coal tar; releases toxic by-products With recombinant DNA E. coli can produce indigo

Transgenic Organisms When recombinant DNA is applied to multicellular organisms, individuals must be bred to yield homozygous individuals Plants may be produced by asexual reproduction (cuttings) Different vectors and gene transfer techniques can be used

Making a Transgenic Plant Figure 19.9 May use Ti plasmids to obtain foreign DNA

Bt Insecticide gene From bacterium Bacillus thuringiensis (bt) Specifies a protein that destroys the stomach lining of certain insect larva 2/3 of U.S. corn is transgenic for the bt gene

Table 19.4

Table 19.5

Transgenic Animals More difficult than plants Several techniques to insert DNA Chemicals to open holes in plasma membrane and liposomes carry DNA in cells Electroporation–a brief jolt of electricity to open membrane Microinjection–uses microscopic needles Particle bombardment – a gun like device shoots metal particles coated with foreign DNA

Figure 19.10

Bioremediation Transgenic organisms can provide process as well as products Ability to detoxify pollutants Examples Hg-contaminated soils GFP gene reveal locations of land mines

Gene Targeting A more precise method for modifying genes of an organism Homologous recombination is a natural process in which DNA sequences, which are similar or identical, recombine (mechanism of crossing over) Introduction of homologous DNA flanking an insertion in a vector allows homologous recombination to occur

Gene Targeting Figure 19.11

Knockout Mutations in Mice May create a model system for studying human disease Identify animal version of a human disease causing allele Transfer the corresponding human mutant allele to mouse ES cells Breed homozygous individuals Use mice to investigate disease symptoms and treatments

Examples of Knockout Mouse Models Severe combined immune deficiency due to ADA gene deficiency Sickle cell disease Polygenic disorders e.g., atherosclerosis by inactivating combinations of lipid metabolism genes Prenatal aspects of neurofibromatosis type I

Normal Knockouts Some genes can be knocked out and have no apparent effect on phenotype Other genes encode the same or similar proteins that can compensate (redundancy) Absent protein does not cause an effect but an abnormal protein might Gene may have no function or different role than hypothesized Conditions required to observe phenotype were not present Example: X collagen in skeletal repair

Monitoring Gene Function Gene Expression Profiling Indicates genes transcribed DNA Variation Screening Detects mutations in Single Gene Polymorphisms (SNPs) Microarray Comparative Genomic Hybridization Deletions and amplifications of DNA sequences between cells or species

Figure 19.12

Figure 19.12

Figure 19.12

Table 19.6

Figure 19.13

Solving a Problem Figure 19.14