Modification of Relative Gene Expression Ratio Obtained from Real Time qPCR with Whole Carotid Body by Using Mathematical Equations

Abstract

Quantitative real time PCR (qPCR) is a common tool used to compare the relative gene expression between treated/untreated cells, different types of tissues, or immature/mature organs. When homogeneous cells are used for qPCR, the Ct number of a tested gene solely represents the quantity of gene expression in cells. However, when a heterogeneous tissue is used for qPCR, the Ct number of a tested gene should be modified depending on several factors: the percentage of each cell type in the sample tissue, the cell type where the target gene is expressed, and the cell type in which the target gene is regulated. The carotid body (CB) is mainly composed of three types of cells: type I (chemoreceptor) cells, type II cells, and other types of cells. Therefore, the relative gene expression ratio obtained from qPCR data using whole CB could be modified by applying one of the following 19 different cases: (1) the target gene is expressed in only one type of cell (3 cases), (2) the gene is expressed in two types of cells and increased in only one or both cell types (9 cases), and (3) the gene is expressed in all three types of cells and increased in only one, two, or all three cell types (7 cases). For example, in the case that the target gene is expressed in all three types of cells and the gene is increased in only a cell comprising 10% of whole CB, the gene expression ratio in that cell will be 9 times as that derived from whole CB. Thus, once the percentage of each cell type in whole CB is observed, the cell type of interest gene (E-gene) expression is identified, and the cell type that regulates E-gene expression by treatment is identified. Thus, the corresponding mathematical equation out of 19 cases could be applied to modify the gene expression ratios measured by qPCR.