Life Science I 83.101.102 Dr. Ekaterina (Kate) Vorotnikova ...
Life Science I 83.101.201 Dr. Ekaterina (Kate) Vorotnikova Office: Olsen 413b E-mail: [email protected] Lecture 26 Genetically modified organisms. DNA profiling. (Page 231; 236-243) GENETICALLY MODIFIED ORGANISMS:
Genetically Modified Bacteria Genetically Modified Plants Genetically Modified Animals Copyright 2009 Pearson Education, Inc. Recombinant cells and organisms can mass-produce gene products Cells and organisms containing cloned genes are used to manufacture large quantities of gene products Capabilities of the host cell are matched to
the characteristics of the desired product Prokaryotic host: E. coli Can produce eukaryotic proteins that do not require posttranslational modification Has many advantages in gene transfer, cell growth, and quantity of protein production Can be engineered to secrete proteins Copyright 2009 Pearson Education, Inc. Eukaryotic hosts Yeast: Saccharomyces cerevisiae Can produce and secrete complex eukaryotic proteins Mammalian cells in culture
Can attach sugars to form glycoproteins Pharm animals Will secrete gene product in milk Copyright 2009 Pearson Education, Inc. Media_26\12_08ATransgenics_DV.mpg Pharm animals that produce a human protein. Examples of products from pharm animals include: goats producing tissue plasminogen activator (TPA) for heart attack victims, sheep producing alpha-1-antitrypsin to treat emphysema, and pigs producing hemoglobin as a blood substitute.
DNA technology has changed the pharmaceutical industry and medicine Products of DNA technology Therapeutic hormones Insulin to treat diabetes Human growth hormone to treat dwarfism Diagnosis and treatment of disease Testing for inherited diseases Detecting infectious agents such as HIV
Copyright 2009 Pearson Education, Inc. Products of DNA technology Vaccines Stimulate an immune response by injecting Protein from the surface of an infectious agent A harmless version of the infectious agent A harmless version of the smallpox virus containing genes from other infectious agents Copyright 2009 Pearson Education, Inc. Equipment used in the production of a vaccine against hepatitis B.
Advantages of recombinant DNA products Identity to human protein Purity Quantity Copyright 2009 Pearson Education, Inc. Genetically modified organisms are transforming agriculture Genetically modified (GM) organisms contain one or more genes introduced by artificial means Transgenic organisms contain at least one gene from
another species GM plants Resistance to herbicides Resistance to pests Improved nutritional profile GM animals Improved qualities Production of proteins or therapeutics Copyright 2009 Pearson Education, Inc. Using the Ti plasmid as a vector for genetically engineering plants. Agrobacterium tumefaciens
DNA containing gene for desired trait 1 Ti plasmid Insertion of gene into plasmid Recombinant Ti plasmid
Restriction site A modified form of the Ti (tumor inducing) plasmid is used in plant genetic engineering. The tumor inducing genes have been removed from the plasmid but genes required for insertion into plant chromosomes have been retained. The gene of interest has been inserted into the plasmid under the control of a bacterial promoter. Agrobacterium tumefaciens Plant cell DNA containing gene for desired trait
1 Ti plasmid Insertion of gene into plasmid Recombinant Ti plasmid 2 Introduction
into plant cells DNA carrying new gene Restriction site Agrobacterium tumefaciens Plant cell DNA containing gene for desired trait 1
Ti plasmid Insertion of gene into plasmid Recombinant Ti plasmid 3 2
Introduction into plant cells Regeneration of plant DNA carrying new gene Restriction site Plant with new trait A mix of golden rice and standard rice.
Gene therapy may someday help treat a variety of diseases Gene therapy aims to treat a disease by supplying a functional allele One possible procedure Clone the functional allele and insert it in a retroviral vector Use the virus to deliver the gene to an affected cell type from the patient, such as a bone marrow cell
Viral DNA will insert the functional allele into the patients chromosome Return the cells to the patient for growth and division SCID (severe combined immune deficiency) was the first disease treated by gene therapy First trial in 1990 was inconclusive Second trial in 2000 led to the development of leukemia in some patients due to the site of gene insertion
Challenges Safe delivery to the area of the body affected by the disease Achieving a long-lasting therapeutic effect Addressing ethical questions Copyright 2009 Pearson Education, Inc. DNA PROFILING Copyright 2009 Pearson Education, Inc. DNA and Crime Scene Investigations DNA evidence was used to solve a double
murder in England Showed that two murders could have been committed by the same person Showed the innocence of someone who confessed to one of the murders Showed the absence of a match in 5,000 men tested when the murderer persuaded another man to donate blood in his name Showed a match with the murderer and DNA found with both victims Media_26\12_15ADNAForensics_DV.mpg Copyright 2009 Pearson Education, Inc.
DNA profiling is the analysis of DNA fragments to determine whether they come from a particular individual Compares genetic markers from noncoding regions that show variation between individuals Involves amplification (copying) of markers for analysis Sizes of amplified fragments are compared The PCR method is used to amplify DNA sequences
Polymerase chain reaction (PCR) is a method of amplifying a specific segment of a DNA molecule Relies upon a pair of primers Short DNA molecules that bind to sequences at each end of the sequence to be copied Used as a starting point for DNA replication Repeated cycle of steps for PCR Sample is heated to separate DNA strands Sample is cooled and primer binds to specific target sequence Target sequence is copied with heat-stable DNA polymerase
Advantages of PCR Can amplify DNA from a small sample Results are obtained rapidly Reaction is highly sensitive, copying only the target sequence Copyright 2009 Pearson Education, Inc. Compare STR (short tandem repeats)
from individuals. Gel electrophoresis separates DNA molecules based on size DNA sample is placed at one end of a porous gel Current is applied and DNA molecules move from the negative electrode toward the positive electrode Shorter DNA fragments move through the gel pores more quickly and travel farther through the gel DNA fragments appear as bands, visualized through staining or detecting radioactivity or fluorescence Each band is a collection of DNA molecules of the same length
DNA profiling has provided evidence in many forensic investigations Forensics Evidence to show guilt or innocence Establishing family relationships Paternity analysis Identification of human remains After tragedies such as the September 11, 2001, attack on the World Trade Center
Species identification Evidence for sale of products from endangered species Copyright 2009 Pearson Education, Inc.
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