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Polymerase Chain Reaction |
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Definition
process to amplify trace amounts of DNA through a cyclical process, useful in Gene cloning, DNA sequencing, Gene expression analysis, DNA fingerprinting, and Detecting infectious and genetic disease disorders. DNA is heated to separate the two strands, then RNA primer is added and dna polymerase synthesizes the two strands: process is repeated as many times as necessary, giving 2^n DNA molecules |
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How do you know if cells are happy |
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morphology, mitotic index (4-5%), plating efficiency and expression of specialized functions (wether they are differentiated or not) |
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single cells, no longer organized as tissues. derived from dispersed cells taken from original tisse |
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derived from an explant tissue, directly from the animal. usually survive for a finite period of time |
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established/continuous cell line |
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a primary culture that has become immortal due to some transformation, most commonly tumor derived or transformed with an oncogen |
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number of successive sub cultures from primary culture |
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A degradative enzyme that recognizes and cuts up DNA (including that of certain phages) that is foreign to a bacterium |
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The direct manipulation of genes for practical purposes |
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DNA in which nucleotide sequences from two different sources are combined in vitro into the same DNA molecules |
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five steps of genetic engineering |
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IITTS Isolation of target gene Insertion into vector Transfer into organism Transform cells in organism Selection |
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To detect the presence or absence of a specific DNA sequence in a sample of DNA using electrophoresis and probe hybridization. |
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It allows us to observe particular gene expression at the level of RNA during differentiation, morphogenesis, and in abnormal or diseased conditions. |
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Used to separate proteins based on size and electrophoretic mobility, Can be used to detect the presence of a specific protein. SDS is mixed with protein, incubated to convert to primary structure and give all negative charge. With current, will move towards positive side through gel |
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Fluorescent Activated Cell Sorting (FACS) |
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Definition
Technique using fluorescent and electrical properties of specific antibodies to separate cells in suspension |
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1) cells are the basic unit of life 2) all organisms are composed of cells 3) cells only arise from other pre-existing |
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1)surrounded by semi-permeable lipid membrane (maintain homeostasis) 2)include DNA (for reproduction and protein synthesis 3)are compartmentalized 4) have same basic mechanisms for energy metabolism 5) organization, react to external stimulation, movement, reproduction |
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rRNA synthesis (no membrane) |
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post translation modification, packages and transports proteins |
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help break down and protect cell from reactive oxygen species i.e. free radicals |
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microtubule organization centers [centrioles] |
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transport already packaged proteins, structure/support, motility, cell division |
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MTs, actin, myosin fibers, intermediary fibers |
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