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摘要In this research,the Cocoonase gene was cloned by RT-PCR as an 860bp fragment,including the signal peptide and the core sequence of Cocoonase gene.In order to investigate the function of signal peptide,recombinant transfer vector pBacPAK8-Cocoonase-EGFP were constructed by fusing with enhanced green fluorescent protein (EGFP) gene to observe under fluorescence microscope.The purified pBacPAKS-Cocoonase-EGFP DNA was co-transfected with linear virus Bm-BacPAK6 DNA into BmN cells.The homologous recombination occurred in the cells and then the recombinant virus Bm-BacPAK8-Cocoonase-EGFP was obtained.BmN cell was infected with the recombinant virus Bm-BacPAKS-Cocoonase-EGFP,and fluorescent signal was detected in most of the cells under fluorescence microscope at 72 h postinfection.Then BmN cells were harvested.Both SDS-PAGE and Western-blotting analysis indicated that the Coooonase was expressed successfully in silkworm(Bombyx mori) baoulovirus expression vector system.Furthermore,refered to Astrup method(Park Y S 1999; Lee S K 2001),used fibrin plate process confirmed that expression product in vitro had cellulolytic activity.We conclude that silkworm expression system can be used successfully to express functional Cocoonase.