Characterization of the double deletion mutant of pseudorabies virus --R23

• Main Authors: Hsu, ching-ming, 許晉銘
• Other Authors: Chang Tien-Jye
• Format: Others
• Language: zh-TW
• Published: 1996
• Online Access: http://ndltd.ncl.edu.tw/handle/00179850513657232064

碩士 === 國立中興大學 === 農業生物科技學研究所 === 84 === Pseudorabies virus （ PRV ）,a pathogen of pseudorabies, is a member ofAlphaherpesvirinae. The major strategy of controlling pseudorabies is todevelop live vaccines of genetically engineered mutants in which their genesof virulence but nonessential in replication were deleted. Inactivation ofThymidine kinase （ TK ） gene leads to a decrease in virulence and latencywithout affecting viral replication. Glycoprotein gE-deleted PRV mutantshows a decrease in virulence and it can be used for differentiat ionbetween wild type virus infection and vaccination. The PRV R23, TK/gEdeletion mutant,is deleted of 400 bp in TK gene and of 1.4kb in gE gene. Thepurpose of this study is to characterize the genomic structure of R23strain. The strains R23 and TNL genomic DNA were characterized by BamHIdigestion and Southern blot hybridization using pTK-1、 pGI-1 DNA as probes.The results showed that the gE gene and TK gene of TNL localized at 7.4 kband 3.1 kb in size respectively while both TK and gE genes of R23 we re atthe same location of 3.1 kb in size. Furthermore, only gE gene of TNL wasexpressed when the anti-gE mAb was used to detect the gE gene expression ofTNL and R23 by western blot analysis. Finally, the results of geneticsequencing demonstrated that the R23 mutant had lost its gE and 11K genes,and the partial gI and 28K genes.