Category Archives: Animal Work

Model Organisms and DNA’s “Molecular Clock”

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“Model organisms” are the best-studied organisms in experimental biology. For a particular research questions a specific “model organism” is chosen for its balance of: (1) ease of use and (2) “generalizability” of results. For example

  • unicellular organisms(e.g. bacteria and yeast): are used to answer questions in basic biochemistry or molecular biology;
  • invertebrates (e.g. worms and flys): are used to answer questions in genetics or embryonic development
  • vertebrates (e.g. zebrafish to primates): are used in models of human disease (as they have requisite physiological and neurological complexity)

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Introduction to RT-PCR (gene/mRNA expression)

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Quantitiative Reverse-Transcriptase PCR (RT-PCR) is different from regular PCR in that it does not measure genes(i.e. DNA) per se but rather measures the EXPRESSION of those gene (i.e. mRNA). Given the fact gene expression (mRNA) can vary dramatically between cell-types, it is important to first isolate a single cell-type by:

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Introduction to PCR and Animal Genotyping

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In a follow up to our overview on DNA methods, we wanted to discuss PCR (polymerase chain reaction) which is one of the most sensitive and versatile techniques in molecular biology. PCR is a technique which selectively amplifies any targeted DNA from a complex mixture based on a set of framing primers. These primers are ~20 base oligonucleotides which we can (1) design based on a sequenced genome and (2) make/order based on solid-phase chemical synthesis. PCR has many applications (see partial list below) but is to test for a particular gene/mutation (i.e. “genotype”) in an animal (see figure above).

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Hybridomas for Large-scale Antibody Production

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Following up on our post introducing antibody-based experiments, we wanted to describe how antibodies are made in large volume: First, an animal is immunized with the protein (or peptide) of interest; Second, the spleen is dissected and the plasma cells producing the antibodies of interested are isolated; Finally, these plasma cells are fused with myeloma cells to “immortalize” them into a hybridoma (an antibody-producing cell-line that can be cultured indefinitely).

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Back-crossing mutant mice into a genetic background

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Inbred mouse lines are used for most laboratory mouse work to make sure that all the mice have nearly identical genetic backgrounds (i.e. genetic code). These lines were originally generated by repeatedly mating mice with their siblings which minimized genetic variability and makes these mice as close to clones of each other as is practically possible.

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Introduction to Mouse Breeding

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Careful breeding/husbandry of mice is important when one is asking genetic questions about disease. Such questions include: “Does gene X contribute to cancer?”, “Does my drug treat cancer by targeting gene X?”.

In general, these questions can only be answered by comparing mice that have gene X (WT or +/+) or don’t have gene X (KO or -/-). To make sure the genetic difference is the only difference between the mice, you usually compare siblings of inbred mouse-lines which have the same sex, age, environment and genetic background.

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