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"I’ve been working with iTL over the past 5 years in the production of 3 different genetically altered mice. Not only did iTL help in the design of the mice, […]” - Raghu Mirmira, MD, PhD University of Chicago.
Creating the ideal flox design for your project can be a difficult task depending on what the end goal might be. Generating conditional knockout mouse models requires a certain degree of precision and thorough research regarding the gene that has to be flanked by loxP sites, as well as the specific methods used for targeting and disrupting the gene through floxing and Cre lox recombination. This ultimately culminates in the process of analyzing the effects of the gene knockout both in the short term and on a long term basis – depending on what the research study requires.
The process of flox design and knocking out genes through the Cre-lox recombination process is fairly simple in theory. Floxing is the process of flanking a gene between two loxP sites in order to target it for knockout. This process of gene targeting is used in ES cells in order to support the development of otherwise healthy mouse models in which a specific, targeted gene is knocked out or deactivated through the use of Cre-lox recombination. Researchers can then study the mouse to find out specifically what the effects of the knockout are, what disorders or developmental issues are caused due to the lack of activity of that particular gene, and why the gene plays a role in creating those issues.
There are numerous benefits to using an effective flox design in order to produce knockout genes. One of them is that genes that could be fatal if deactivated can be safely analyzed without any risks to human patients. The mice that take part in the experiment are genetically similar enough to humans in order to ensure that scientists are able to come up with viable results that may ultimately be put to use in creating drugs and treatments.
Once the flox design is obtained by flanking a gene segment between two loxP sites, the process of Cre-mediated recombination is somewhat more complex in practice than in theory. It involves conditional mutant mice that are obtained through crossing the floxed strain with a transgenic line. This action has the purpose of generating the knockout in vivo with the use of Cre. What makes the process easier and more manageable is the fact that there have been many transgenic lines generated over the years. Today it’s very easy to find the correct line that would support most studies based on tissue-specific conditional knockouts.
The Cre-mediated recombination process combined with floxing can lead to excellent results, whether the purpose is to study the development of cancerous tumors in specific tissues or the progression of genetic diseases that are rare, but can be quite dangerous or even fatal. Proper flox design and Cre-lox recombination research are an important part of developing treatments that could help people lead better and healthier lives in the near future.
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