The Establishment and Application of Persistent Infection Using Baculovirus Autographa californica Nuclear Polyhedrosis Virus

• Main Authors: Lai Kuei-Tai, 賴癸太
• Other Authors: Chiang Ann-Shyn
• Format: Others
• Language: zh-TW
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/18311843381689652240

碩士 === 國立清華大學 === 生命科學系 === 87 === Abstract Persistent viral infection in insect cells is a prominent though poorly understood phenomenon. In these studies, we have successfully established persistent viral infected cell lines and created a new baculovirus expression system for foreign protein production. Clones of surviving Spodoptera frugiperda (Sf) cells harboring persistent viral genomes were derived from the infection of baculovirus from the infection of baculovirus AcMNPV carrying a deletion at the apoptosis supresssor gene p35 or a temperature sensitive mutation in late expression factor 4 (lef-4) gene. The viral genomes were detected after long passages in these persistent infected cells and were reactivated by transfection of p35 for the former or by reculturing under permissive temperature for the latter cases. Continuous release of high titer viruses was evident in some of the persistent viral infected cells resulted from the infection of AcMNPV with p35 deletion. Interestingly, the released viral progeny could not establish persistent viral infection. The persistent viral infected cells resulted from p35 deletion became highly resistance to super infection and suppress the expression of late (vp39) and very late (p10) genes. By contrary, the persistent viral infected cells established by lef-4 mutant of AcMNPV released low viral titers and these viral progenies could establish persistent viral infection at nonpermissive temperature. The expression of late (vp39) and very late (p10) genes were normal during productive viral infection. However, due to an abnormal lef-4 function at nonpermissive temperature these promoters were highly suppressed during persistent viral infection. These results and phenomenon might be suggested for further experiments about the mechanisms of persistent viral infection and virus resistance. In this study, EGFP was also used as a reporter gene to monitor the expression of foreign gene in these two persistent viral infection systems.