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36 Cards in this Set
- Front
- Back
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What are the deleterious effects of DNA homologous recombination?
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deletions, inversions, translocations and rearrangements leading to some cancers
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What are the vectors for recombination technology?
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1) PLASMIDS - to clone short fragments
2) PHAGES - to clone <20kb 3) YEAST - to clone million of bps |
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What are restriction endonucleases?
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Recognize specific 4-, 6- and 8-base pair sequences (paladromes) and make restriction fragments with cohesive ends. Ligase joins the two together to make a recombinant molecule
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What is a cDNA library?
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a collection of clones (vector + DNA/cDNA) of whole genome fragments of cDNAs corresponding to mRNA fragments
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How can mRNA be isolated?
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by taking advantage of Poly A tails
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What is the process of hybridization?
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denature by increasing temperature, renature and allow complementary strands to anneal. Probe hybridization to find specific sequence
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What is Northern blotting used for and how is it done?
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RNA Analysis. mRNA is separated, transferred to a membrane and probe hybridized.
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What is Southern blotting used for and how is it done?
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DNA Analysis. Restriction digestion, electrophoresis, transfer DNA to membrane and use with probe hybridization.
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What is in situ hybridization?
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Chromsomal location can be determined by making a map by hybridizing probe fragments
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What is Western blotting used for?
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Separation of proteins by electrophoresis by 2D or 1D separation. Then they are transferred to a membrane and ID by antibody binding (antibody detection by color development)
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How many base pairs are there in the human genome?
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3.1647 billion base pairs
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How many genes are there in the human genome?
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~25,000
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Where are functional genes found in the human genome?
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G-C rich regions
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What are the differences between the human nuclear genome and mitochondrial genome?
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Nuclear genome contains 24 chromosomes, many more base pairs and genes but only 1.5% is coding.
Mitochondrial genome is only 1 circular double strand with 16.6 kb and 37 genes. But ~93% is coding. |
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What are the biological roles of homologous recombination?
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Genetic assortment in gametogensis, DNA repair and clearing replication hurdles
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What is a Holliday Junction?
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produced by reciprocal exchange of strands between two DNA double helices
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What catalyzes the synapse step in general recombination?
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RecA type of protein bound to a DNA single strand
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What is the genetic cause for chronic myeloid leukemia
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small philadelphia chormosome.
ABL oncogene on chromosome 9 is joined with 5' part of BCR gene on chromosome 22 (via translocation) |
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What is the genetic cause for Burkitt's Lymphoma?
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MYC oncogene from chromosome 8 is translocated to IGH locus on chromosome 14
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What is positional cloning?
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Identifying or cloning a disease gene based on its chromosomal location
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How do you find chromosome location?
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linkage mapping. Linking a disease region using polymorphic markers. Analyze the region for the presence of likely candidate and confirm that the candidate gene is mutated in affected individuals.
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What is the genetic and phenotypic traits of neurofibromatosis 1?
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autosomal dominant disease, linked to chromosome 17
causes brown spots on skin, benign nodules on iris, neurofibromas and growth abnormalities/deformities |
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What is an LOD score?
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The logarithm of ratio of likelihoods.
The likelihoods of a linkage between two loci at a specific recombination fraction |
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How can gene therapy correct defects in somatic cells?
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Gene augmentation, gene replacement, targeted inhibition and targeted killing
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How does gene augmentation gene therapy work?
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introduce a functional copy into the body to treat a loss-of-function disease
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How does gene replacement (in situ) correction work?
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used for gain of function diseases
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What does targeted inhibition gene therapy do?
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inhibits specific activate oncogene or pathogens
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What is an example of targed killing gene therapy?
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cancer treatments
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What are two approaches to gene therapy?
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1) In vivo (directly injecting cloned gene)
2) Ex vivo (removing cells and correcting them outside the body, then putting them back in) |
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What are the delivery methods to get genes into cells?
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Viruses and nonviral
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What are the viral methods of delivering genes into cells?
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adenovirus, adeno-associated virus, lentivirus and HSV
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What are the nonviral methods of delivering genes into cells?
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Liposomes, injection/bombardment, receptor mediated
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What happens in homologous recombination gene therapy?
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a mutant gene can be replaced by a normal gene
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Wha happens in antisense oligonucleotide gene therapy?
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a disease causing gene can be inactivated
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What happens in Rybozyme gene therapy?
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the mRNA transcript of a disease causing gene is degraded
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What happens in RNA interference gene therapy?
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the mRNA of the target gene is targeted for degradation
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