Knockout Mouse Technology: A Genetic Analysis

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Knockout mouse technology is valuable for determining the purposes and operations of genes that have not been sufficiently studied in the laboratory. These genes are replaced with mutant or diseased versions of themselves, permitting researchers to discover and discern how losing the normal function of certain genes can affect an organism. Through the use of this technology, the mouse genome has been extensively mapped and attributed to a variety of functions for specific genes. In this analysis, I will discuss knockout mouse technology, describe the systematic process used in the lab, and introduce the concept of conditional knockout mouse technology pertaining to Cre (bacteria) recombination. In order to begin the process of manipulating …show more content…
Hypothetically, assume that a researcher has a white, pregnant female mouse that has a gene with an unknown function. In order to mutate this gene, the researcher uses a culture growth of embryonic stem cells from a different color mouse (agouti color) and induces the specific genetic mutation via synthetically modified DNA. This DNA includes components required to test the embryonic stem cells via positive and negative selection for the desired mutation. Positive selection involves the use of G418 (antibiotic), whereas negative selection involves the use of ganciclovir (drug that combats viruses). The DNA inserted into the embryonic stem cells was composed of two indicator genes: neor, which is resistant to G418, and tkHSV, which is degraded and destroyed by ganciclovir. The researcher wants to keep the genes that are resistant to G418 (neor) because this means that homologous or nonhomologous recombination has occurred. Thus, the cells that did not undergo a form of recombination are destroyed; however, the researcher wishes to only maintain cells with genes …show more content…
Conditional knockout mouse technology allows researchers to select and mutate a gene at different points throughout the developmental process or a gene of certain category of cells. This is important because it allows researchers to study how different genes are important at separate times and locations throughout development, which also allows researchers to determine the functions of these particular genes. Because it is difficult for the researchers to know exactly when and where the gene is important during development, they must come up with a method to get around mouse and/or cellular death. Cre recombination allows researchers to manipulate the DNA of an organism. Cre is an enzyme that drives recombination of the DNA. The desired gene’s DNA should be identified in the same way as the previously described knockout mouse technology process, and this should result in isolation of DNA that has undergone homologous recombination. This conditional knockout process will involve two mice: a loxP mouse and a Cre mouse. After homologous recombination in the loxP mouse, two loxP sites will flag the exon of the desired gene. Another mouse, termed the Cre mouse, contains the promoter gene coding for the Cre enzyme (note that this Cre condition was created with the use of embryonic stem cells). If the loxP mouse and the Cre mouse are mated, then

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