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46 Cards in this Set
- Front
- Back
- 3rd side (hint)
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Where in the body can adult stem cells be found?
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Umbillical cord
Bone Marrow Liposuction fat Other tissue |
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Where can embryonic stem cells be found?
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In aborted fetuses/IVF embryos
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What is good about ESC that ASC don't have?
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The zygotes are totipotent and the cells on the interior of the blastocyst are pluripotent, so they are much more versatile than differentiated cells.
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What are the steps in therapeutic cloning?
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1. Haploid egg is removed from 1st person; the nucleus is removed.
2. Epithelial cells are removed from the 2nd person; cells are deprived of nutrients and stop dividing. 3. Diploid nucleus of epithelial cell replaces haploid nucleus of egg. 4. Cell is shocked with an electric charge. 5. Egg grows, divides like normal zygote. |
The point is to a human embryo with the goal of harvesting the stem cells in them.
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What are the benefits of therapeutic cloning?
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You don't have to use a stem cell; you can create one and avoid controversy.
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What are the benefits of stem cell treatment?
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Replace lost tissue
No risk of rejection |
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What is an IPS?
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Induced Pluripotent Cell
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How are IPS' used?
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Skins cells in mice with Sickle Cell Anemia are reprogrammed into ESC-like cells. The mutation causing the disease is corrected and differentiated into blood stem cells, at which point they are transplanted.
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What is the main challenge of stem cells?
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Making them differentiate into the type of cell you want.
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What strategy is used to get stem cells to differentiate into the type of cell you want?
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Place them in the right part of the body and hope they differentiate correctly.
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What is useful about a retroviral vector?
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Viruses have special molecular mechanisms to efficiently transport genomes inside the cells they infect. Retroviral vectors harness this technology to transduct genes into cells.
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How are vectors used in gene therapy?
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To deliver unmutated copies of genes.
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What problems occur when using retroviral vectors?
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1.The body can experience an immune response that sends the patient into septic shock and kills him (Jesse Gelsinger, 1999)
2. The gene may be inserted in a location that can cause diseases (XSCID trial 2002, two children got leukemia). |
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What are the steps of gene therapy using retroviral vectors?
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1. Insert RNA of normal allele into retrovirus
2. Let retrovirus infect bone marrow cells that have been removed from the patient and cultured. 3. Viral DNA carrying the normal allele is inserted into chromosome 4. Inject engineered cells into the patient. |
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What are three examples of problems gene therapy can be used to fix?
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1. Genetic disorders: normal version of mutant gene reintroduced.
2. Autoimmune disorders: give immune modulating gene 3. Cancer: gene kills cells, improves immune function, and makes cancer cells more sensitive to chemo radiation. |
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What is SCID?
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Severe combined immunological deficiency (malfunction in ADA gene)
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Name four problems associated with gene therapy
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-Short lived
-Immune response can be overwhelming -Multi gene disorders hard to treat with viral vectors -Need human guinea pigs |
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What is a stem cell?
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A single cell that can replicate itself or differentiate into many cell types.
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What are the two types of stem cell?
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Embryonic: from zygotes (totipotent cells) or the insides of blastocysts (pluripotent)
Adult: differentiated (pluripotent cells) maintained in our bodies to replace old/dead cells. |
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How are stem cells cultivated?
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1. In vitro fertilized eggs
2. When blastocyst is 5-7 days old, harvest its inner stem cell mast 3. Culture undifferentiated stem cells. |
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What is a retrovirus?
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A virus that injects RNA rather than DNA into its host with instructions to create DNA from RNA using reverse transcriptase.
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Where are retroviral vectors injected in the body?
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In the part of the body where it is needed. (Retnal Blastoma: in the eye)
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What are the problems with gene therapy?
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The gene can be injected into a vital part of the genome that will disrupt it or even cause the cell to replicate uncontrollably (cancer)
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What is a DNA microarray?
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A computer chip with 25 base pair probes. Samples of infected and noninfected tissue are applied to the array together; DNA from the samples that finds its cDNA complement on a probe is illuminated.
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What is BT?
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A bacteria in soil that contains a gene toxic to bugs. When put in corn genes, bugs do not eat the corn.
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When was BT corn approved?
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1996
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What controversy surrounds it?
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Scientists worried that BT corn might cross pollinate with regular corn and kill monarch butterflies, but in 2002 it was definitively determined that this isn't the case.
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What are the four types of genetic variation in bacteria?
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Mutation
Transformation Transduction Conjugation |
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What is an R plasmid?
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A plasmid containing transposons with resistance genes
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What is an F plasmid?
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A bacterial DNA sequence that allows bacteria to produce sex pili needed for conjucation.
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What does In Vivo gene therapy entail?
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In Vivo gene therapy does not remove cells from the patient's body; vectors are used to deliver the DNA to the required location.
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What does Ex Vivo gene therapy entail?
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Cells are removed from the patient's blood or bone marrow and cultured in a lab. They are then infected with a virus carrying the normal gene that is inserted into the cells that were removed. The cells with the recombinant DNA are then reinjected back into the patient.
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What are four methods for vector gene delivery>
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Viral vector (must be treated first so as not to be pathogenic)
Naked DNA (not very effective) Liposomes: hollow microspheres stuffed with DNA DNA gun |
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What is transformation?
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A process by which vectors are taken up by bacteria with the help of chemicals or electric shock to make the bacteria cell walls porous.
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What is the point of cDNA?
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cDNA is DNA derived from the mRNA of a normal cell for use in vectors, etc. It is formed by adding reverse transcriptase to the mRNA for the gene.
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What is cDNA?
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DNA that is synthesized from processed mRNA, where introns have already been excised. cDNA is important because prokaryotes used to produce eukaryotic proteins do not always have the tools to process pre mRNA.
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How do you make cDNA?
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Isolate the mRNA from cells and add revers transcriptase to make the first strand.
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How does the DNA polymerase know where to start synthesizing?
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Because of the poly-A tail added at the 3' end. It then loops back around in a hairpin loop to create the second strand. The loop is eventually removed by an s1 nuclease.
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What is so great about bacteria?
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-They can grow where no human could
-The can produce anything from plastic to human proteins. -They clean up our waste, make up our food -They are a bigger part of us than human cells |
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Define transformation.
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DNA from a dead S cell leaks out and replaces the DNA of a live R cell. The new DNA exchanges with a similar sequence in a bacterial chromosome; the R cell acquires some S cell traits.
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Define transduction.
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A phage infects a bacterial cell and the host DNA is hydrolyzed into pieces. The phage DNA and proteins are made. Occasionally a bacterial DNA fragment is packaged in a phage capsid. Transducting phages infect new host cells where recombinantion occurs.
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What are the steps for gene cloning (includes transformation)
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1. Cut a plasmid with restriction enzymes at the indicated site.
2. Cut out sample gene with the same enzymes; test to make sure gene is correct by running a gel. 3. Ligate the gene into the plasmid 4. Help bacteria transform the recombinant plasmid by shocking it or applying heat. 5. Test to see which bacteria have the plasmid by growing them on antibiotic culture. 6. Test to make sure bacteria have the right gene by applying original restriction enzymes and running a gel for size. 7. Culture the correct bacteria. |
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How is cDNA made?
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1. Isolate mRNA
2. Use reverse transcriptase to make 1st strand. 3. Degrade mRNA 4. Make 2nd strand. |
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What is a DNA library?
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A collection of cloned DNA that are derived from restriction fragments.
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How are they made?
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1. Cut up the whole genome with restriction enzymes.
2. Clone the fragments into a vector. 3. That collection of cloned DNA is a library. |
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How do you find one particular gene in a library?
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By using a probe.
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