Generation Of A Dual Labeling System For Insulin Positive And Negative Cells

Cell labeling and tracing provides an indispensable approach to studying pancreas development and regeneration. However, the traditional Cre-LoxP system consists of two vectors, and was generally used for tracing a single cell population during development. In this study, we proposed a single vector system with a dual labeling capacity for simultaneous tracing of insulin positive and negative cells. Surprisingly, we found that the tissue specificity of RIP was significantly reduced due to the enzymatic amplification of Cre. In order to improve tissue specific labeling, we determined to lower the enzymatic activity of Cre by reducing its expression levels, by generating a two-hour degradation version, or by introducing point mutations. The results showed that an R173H point mutation in Cre maintained efficient recombination in beta cell line NIT-1 when controlled by RIP, while undetectable activity was found in Ad293 cells. This vector could be an invaluable tool in studying islet biology and pancreas regeneration.