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Course: MCAT > Unit 2

Lesson 1: Foundation 1: Biomolecules

DNA technology: Genetic recombination and conditional knockouts


A "conditional knockout" defines an animal model in which a gene of interest is either: (1) inactivated only in specific cell types such that other cell types exhibit unmodified, functional gene expression (tissue-specific knockout model); or (2) temporarily suppressed at a given time-point in development (inducible knockout model).
Researchers can use enzymes such as Cre recombinase to engineer tissue-specific knockouts. Cre recombinase cleaves DNA at palindromic consensus sequences called loxP sites. If researchers synthesize a gene flanked by loxP sites (called a “floxed” locus), this locus is susceptible to elimination in the presence of Cre recombinase, as shown in Figure 1. A key step in generating conditional knockouts is selecting a promoter that will drive the expression of the inserted Cre gene coding for CRE recombinase but only in certain cell types.
Figure 1 Cre-Lox Conditional Knockout Technology
Researchers used this technology to examine the effects of myeloid-specific Caveolin 1 (CAV1) knockouts. CAV1 is a ubiquitously expressed gene that codes for a protein involved in endocytosis and phagocytosis. Researchers engineered mice with floxed CAV1 alleles and used a LysM promoter to express Cre recombinase. Myeloid cells (including monocytes, macrophages, and neutrophils) express LysM, the gene coding for lysozyme M.
Figure 2 LysM promoter specifies Cre expression (CDS = coding sequence)
Rank mice of the following genotypes in order of decreasing vulnerability to bacterial infection.
I. Heterozygous for LysMCre; CAV1flox/flox
II. Homozygous for LysMCre; CAV1+/–
III. Wild-type LysM; CAV1+/+
IV. Heterozygous for LysMCre; CAV1 flox/–
Choose 1 answer: