Generation of General and Tissue-Specific Gene Knockout Mouse Models

Knockout technology has established the functions of many genes affecting plasma lipid and lipoprotein levels and the development of atherosclerosis. However, many genes remain to be characterized. The ability to produce mice lacking whole-body expression of a given gene is still one of the most powerful techniques available for determining gene function. A complementary approach, underutilized yet vitally important to understanding lipoprotein metabolism, is the ability to create mice with gene deficiency only in a specific tissue. Liver, intestine, and macrophages are the major tissues and cells involved in lipoprotein metabolism and atherosclerosis, and additional tissues such as adipose tissue and brain are also of interest. Thus, feasible approaches to prepare general and tissue-specific gene knockout mouse models are necessary. Here, we describe our general procedure for generating whole-body knockout mice, using as an example the preparation of general (whole-body) phospholipid transfer protein (PLTP) gene knockout mice. We also describe several approaches to generating liver, intestine, and myeloid cell-specific tissue-specific knockout mice, using as an example the preparation of tissue-specific knockout mice for the subunit 2 of serine palmitoyltransferase (SPT), a key enzyme for sphingomyelin de novo synthesis. Bone marrow transplantation is an alternative means of creating myeloid cell-specific knockout mice. The general principles and techniques described here apply to the establishment of other gene knockout mouse models as well. The ability to manipulate gene expression in specific tissues as well as throughout the entire body of the mouse is anticipated to yield novel insights into lipid and lipoprotein metabolism and the development of atherosclerosis.