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    • 2. 发明申请
    • ALLELIC EXCHANGE MUTAGENESIS IN MAP
    • 地图中的等位交换致突变
    • WO2009009798A3
    • 2009-07-30
    • PCT/US2008069999
    • 2008-07-14
    • UNIV WASHINGTON STATEDAVIS WILLIAM CHAMILTON MARY JODAHL JOHNPARK KUN TAEK
    • DAVIS WILLIAM CHAMILTON MARY JODAHL JOHNPARK KUN TAEK
    • C12N15/31
    • C12N15/102A61K39/04A61K2039/522C12R1/325
    • Particular aspects provide efficient allelic exchange methods to generate directed mutations within genes of slow-growing stains of mycobacteria (e.g., Mycobacterium avium subsp. paratuberculosis (Map), Map10 or GFP-expressing Map K-10) using a phage-delivery system, and demonstrate high efficiency allelic exchange. Additional exemplary aspects provide non-naturally occurring slow-growing strains of mycobacteria (e.g., Map, M. bovis, M. tuberculosis) having at least one gene deletion (e.g., pknG, relA, lsr2, panC, panD, proC, trpD, sapM (MAP3432), lysA_1, leuD, and leuC, and deletion mutants at the orthologous loci of two known virulence genes in pathogenic mycobacteria (relA and pknG) and one gene related to colony morphology and biofilm formation in fast growing mycobacteria (lsr2) were made. Further aspects provide novel vaccines comprising such deletion mutants, or portions thereof, and methods for making said vaccines. Yet further aspects provide methods for protecting a mammal from virulent Map, M. bovis, or M. tuberculosis, comprising treating the mammal with the inventive vaccines.
    • 具体方面提供了使用噬菌体递送系统在缓慢生长的分枝杆菌染色体(例如鸟分枝杆菌副结核亚种(Map),Map10或GFP表达Map K-10)的基因内产生定向突变的有效等位基因交换方法,以及 展示高效率的等位基因交换。 其他示例性方面提供了具有至少一个基因缺失的非天然存在的慢性分枝杆菌菌株(例如,Map,M.bovis,结核分枝杆菌)(例如pknG,relA,lsr2,panC,panD,proC,trpD, 在致病性分枝杆菌中两种已知毒力基因(relA和pknG)的直系同源基因座上的sapM(MAP3432),lysA_1,leuD和leuC以及与快速生长的分枝杆菌(lsr2)中的菌落形态和生物膜形成有关的一种基因的缺失突变体分别是 进一步的方面提供了用于保护哺乳动物免受毒性Map,牛分枝杆菌或结核分枝杆菌的方法,所述方法包括用 本发明的疫苗。