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MCAT

Unit 2: Lesson 1

Foundation 1: Biomolecules

Genetic recombination and conditional knockouts

Problem

A "conditional knockout" defines an animal model in which a gene of interest is either inactivated in specific cell types such that other cell types exhibit unmodified, functional gene expression, (Tissue-specific knockout model), or temporarily suppressed at a given time-point in development (Inducible knockout model).
Researchers can use enzymes such as Cre recombinase to engineer tissue-specific knockout mice. Cre recombinase cleaves DNA at consensus sequences called loxP sites. If researchers synthesize a gene flanked by loxP sites (this would be called a “floxed” locus), that locus is susceptible to elimination in the presence of Cre recombinase (Figure 1).
Figure 1 Cre-Lox Conditional Knockout Technology
Caveolin-1 (Cav-1) is a ubiquitously-expressed gene that codes for a protein that is important for endocytosis and phagocytosis. Although a total body knockout (Cav-1 -/-) is nonlethal, researchers are interested in examining the the effects of a myeloid-specific Cav-1 knockout. A key step in generating this conditional knockout is selecting a promoter to drive expression of the inserted Cre gene. For example, myeloid cells (including monocytes, macrophages, and neutrophils) express LysM, the gene for lysozyme M. Using the LysM promoter, one can express Cre recombinase in these myeloid cells only (Figure 2).
Figure 2 LysM promoter specifies Cre expression (CDS = coding sequence)
What percent of the offspring of a male heterozygous for both LysMCre (LysMCre/wt) and Cav-1 (Cav-1–/+) and a female wild type LysM (LysMwt/wt) and floxed, heterozygous Cav-1 (Cav-1-/flox) would have greater Cav-1 expression than either parent?
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