Structure Analysis of Mouse Gene Encoding Kininogens, and Construction of a Vector for Targeted Disruption of Kininogen Gene.

点击翻译按钮获取中文摘要

Mammalian expresses two types of protein precursor for kinins termed high-molecular-weight and low-molecular-weight kininogens which are coded by a single gene and produced by alternative splicing of the gene transcript. To clarify roles of the kallikrein-kinin system in the physiological and pathological conditions, steps have been taken toward the generation of mice with a "knockout" of the kininogen gene. A genomic clones of the mouse kininogen were isolated from129 strain mice using LA-PCR method and their coding sequences determed by DNA sequence analysis. A physical map of the DNA flanking this coding sequence was generated. According to the structure of kininogen gene, a vector was constructed for targeted disruption of the mouse kininogen gene in plasmid pPNT. This vector contains 7.5 Kb of DNA upstream from the bradykinin coding sequence, a neomycin resistance gene, and 2.4 Kb of DNA downstream from bradykinin coding sequence. Thus, the correct homologous recombination event will result in a chromosome in which the coding sequence for the bradykinin in mouse kininogen is replaced with the neomycin resistance gene. This targetting vector has been transfected into embryonic stem cells, and clones containing a targeted disruption of the mouse kininogen will be identified.