Cellulase Gene Clone, Genetic Transformation And The Research On Cre/loxP System

A large number of cellulose become essential in the lives of human being with the use of cellulose enzymes. It has great importance to solving the world's energy crisis, food shortages and environmental pollution, etc. In addition, bio-security issues have gradually become a topic of global concern with the development of modern biotechnology, especially recombinant DNA technology and the genetic engineering. Deleting the marker gene (herbicide-resistant gene) using Cre/loxP system is vital to bio-security issues. In this paper, the specific work as follows:1. Using RT-PCR method, endo-glucanase gene (EGâ…¡) and exo-glucanase gene (CBHâ…¡) have been cloned from the green trichoderma. Exo glucanase gene (CBHâ…¡) has three introns and four exons, while the endoglucanase gene (EGâ…¡) no intron.2. Through adding endoplasmic reticulum stranded signal in EGâ…¡gene, a P3300-35S-EGâ…¡-Tnos vector is built and then transferred to tobacco. 7 transgenic plants have been identified by PCR.3. Using the Cre/loxP system and the male sterility gene to build expression vectors, delete the marker gene (herbicide-resistant gene) and transfer plant expression vector P3300 Cre/lox-Osg6B-Barnase-Tnos into bent grass. 6 transgenic plants pass though PCR identification.