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    Recombinant negative strand RNA virus
    2.
    发明授权
    Recombinant negative strand RNA virus 失效
    重组负链RNA病毒

    公开(公告)号：US5578473A

    公开(公告)日：1996-11-26

    申请号：US209178

    申请日：1994-03-10

    申请人： Peter Palese ,  Jeffrey D. Parvin ,  Mark Krystal

    发明人： Peter Palese ,  Jeffrey D. Parvin ,  Mark Krystal

    IPC分类号： A61K39/00 ,  A61K39/02 ,  A61K39/12 ,  A61K39/13 ,  A61K39/21 ,  A61K39/245 ,  A61K39/29 ,  C07K14/00 ,  C07K14/11 ,  C07K14/16 ,  C07K14/705 ,  C07K19/00 ,  C12N7/00 ,  C12N9/12 ,  C12N9/24 ,  C12N15/09 ,  C12N15/10 ,  C12N15/11 ,  C12N15/86 ,  C12P21/02 ,  C12R1/91 ,  C12N7/01 ,  C12N7/02 ,  C12N7/04

    CPC分类号： C07K14/005 ,  C12N15/10 ,  C12N15/11 ,  C12N15/86 ,  C12N9/127 ,  C12N9/2402 ,  C12Y302/01018 ,  A61K2039/5256 ,  A61K39/00 ,  C12N2740/16122 ,  C12N2740/16222 ,  C12N2760/16022 ,  C12N2760/16122 ,  C12N2760/16143 ,  C12N2840/203 ,  C12N2840/85

    摘要： Recombinant negative strand virus RNA templates which may be used to express heterologous gene products and/or to construct chimeric viruses are described. Influenza viral polymerase, which was prepared depleted of viral RNA, was used to copy small RNA templates prepared from plasmid-encoded sequences. Template constructions containing only the 3' end of genomic RNA were shown to be efficiently copied, indicative that the promoter lay solely within the 15 nucleotide 3' terminus. Sequences not specific for the influenza vital termini were not copied, and, surprisingly, RNAs containing termini identical to those from plus sense cRNA were copied at low levels. The specificity for recognition of the virus-sense promoter was further defined by site-specific mutagenesis. It was also found that increased levels of vital protein were required in order to catalyze both the cap-endonuclease primed and primer-free RNA synthesis from these model templates as well as from genomic length RNAs. This indicated that this reconstituted system had catalytic properties very smilar to those of native viral RNPs. High levels of expression of a heterologous gene was obtained using the constructs and methods described.

    摘要翻译： 描述了可用于表达异源基因产物和/或构建嵌合病毒的重组负链病毒RNA模板。 用于制备耗尽病毒RNA的流感病毒聚合酶用于复制由质粒编码序列制备的小RNA模板。 显示仅含有3'末端基因组RNA的模板结构被有效拷贝，表明启动子仅位于15个核苷酸3'末端内。 不特定流感重要终点的序列没有被复制，令人惊讶的是，含有与正义cRNA相同的末端的RNA被拷贝到低水平。 通过位点特异性诱变进一步定义识别病毒感应启动子的特异性。 还发现需要增加水平的重要蛋白质以便从这些模型模板以及来自基因组长度RNA的催化端帽内切核酸酶引物和无引物的RNA合成两者。 这表明这种重构系统具有与天然病毒RNP非常相似的催化性质。 使用描述的构建体和方法获得异源基因的高水平表达。

    Recombinant negative strand RNA virus expression-systems
    3.
    发明授权
    Recombinant negative strand RNA virus expression-systems 失效
    重组负反馈RNA病毒表达系统

    公开(公告)号：US5166057A

    公开(公告)日：1992-11-24

    申请号：US527237

    申请日：1990-05-22

    申请人： Peter Palese ,  Jeffrey D. Parvin ,  Mark Krystal

    发明人： Peter Palese ,  Jeffrey D. Parvin ,  Mark Krystal

    IPC分类号： A61K39/00 ,  A61K39/02 ,  A61K39/12 ,  A61K39/13 ,  A61K39/21 ,  A61K39/245 ,  A61K39/29 ,  C07K14/00 ,  C07K14/11 ,  C07K14/16 ,  C07K14/705 ,  C07K19/00 ,  C12N7/00 ,  C12N9/12 ,  C12N9/24 ,  C12N15/09 ,  C12N15/10 ,  C12N15/11 ,  C12N15/86 ,  C12P21/02 ,  C12R1/91

    CPC分类号： C07K14/005 ,  C12N15/10 ,  C12N15/11 ,  C12N15/86 ,  C12N9/127 ,  C12N9/2402 ,  C12Y302/01018 ,  A61K2039/5256 ,  A61K39/00 ,  C12N2740/16122 ,  C12N2740/16222 ,  C12N2760/16022 ,  C12N2760/16122 ,  C12N2760/16143 ,  C12N2840/203 ,  C12N2840/85

    摘要： Recombinant negative strand virus RNA templates which may be used to express heterologous gene products and/or to construct chimeric viruses are described. Influenza viral polymerase, which was prepared depleted of viral RNA, was used to copy small RNA templates prepared from plasmid-encoded sequences. Template constructions containing only the 3' end of genomic RNA were shown to be efficiently copied, indicative that the promoter lay solely within the 15 nucleotide 3' terminus. Sequences not specific for the influenza viral termini were not copied, and, surprisingly, RNAs containing termini identical to those from plus sense cRNA were copied at low levels. The specificity for recognition of the virus-sense promoter was further defined by site-specific mutagenesis. It was also found that increased levels of viral protein were required in order to catalyze both the cap-endonuclease primed and primer-free RNA synthesis from these model templates as well as from genomic lengths RNAs. This indicated that this reconstituted system had catalytic properties very similar to those of native viral RNPs. High levels of expression of a heterologous gene was obtained using the constructs and methods described.

    摘要翻译： 描述了可用于表达异源基因产物和/或构建嵌合病毒的重组负链病毒RNA模板。 用于制备耗尽病毒RNA的流感病毒聚合酶用于复制由质粒编码序列制备的小RNA模板。 显示仅含有3'末端基因组RNA的模板结构被有效拷贝，表明启动子仅位于15个核苷酸3'末端内。 不具体针对流感病毒末端的序列没有复制，令人惊讶的是，含有与正义cRNA相同的末端的RNA被复制在低水平。 通过位点特异性诱变进一步定义识别病毒感应启动子的特异性。 还发现需要增加水平的病毒蛋白以便催化来自这些模型模板的帽内切核酸酶引物和无引物的RNA合成以及来自基因组长度的RNA。 这表明这种重构系统具有与天然病毒RNP非常相似的催化性质。 使用描述的构建体和方法获得异源基因的高水平表达。

    Saccharomyces cerevisiae expressing M.sub.2 protein of influenza A virus
    4.
    发明授权
    Saccharomyces cerevisiae expressing M.sub.2 protein of influenza A virus 失效
    表达甲型流感病毒的M2蛋白的酿酒酵母（Saccharomyces cerevisiae）

    公开(公告)号：US5691189A

    公开(公告)日：1997-11-25

    申请号：US556124

    申请日：1995-11-09

    申请人： Stephen E. Kurtz ,  Mark Krystal

    发明人： Stephen E. Kurtz ,  Mark Krystal

    IPC分类号： C07K14/11 ,  C12N1/15 ,  C12N15/81 ,  C12N1/19

    CPC分类号： C07K14/005 ,  C12N15/81 ,  C12N2760/16122

    摘要： A modified Saccharomyces cerevisiae cell, wherein the cell expresses the Influenza virus ion channel protein M.sub.2. Also disclosed is a process for detecting modulators of M.sub.2, which comprises (a) treating such modified Saccharomyces cerevisiae cells with a test substance, and (b) assessing growth in the presence of a test substance, wherein a change in growth signals that the test substance is an M.sub.2 modulator. M.sub.2 inhibitors are useful anti-viral agents.

    摘要翻译： 一种修饰的酿酒酵母细胞，其中所述细胞表达流感病毒离子通道蛋白M2。 还公开了一种用于检测M2的调节剂的方法，其包括（a）用测试物质处理这种修饰的酿酒酵母细胞，和（b）评估测试物质存在下的生长，其中生长信号的变化表明测试 物质是M2调制剂。 M2抑制剂是有用的抗病毒剂。

    Capped nucleic acid oligomers that inhibit cap-dependent transcription of the influenza virus endonuclease
    5.
    发明授权
    Capped nucleic acid oligomers that inhibit cap-dependent transcription of the influenza virus endonuclease 失效
    抑制流感病毒核酸内切酶转录的封闭的核酸低聚物

    公开(公告)号：US5837852A

    公开(公告)日：1998-11-17

    申请号：US136214

    申请日：1993-10-14

    申请人： Thomas D. Y. Chung ,  Christopher W. Cianci ,  Moira Hagen ,  Mark Krystal ,  Richard J. Colonno

    发明人： Thomas D. Y. Chung ,  Christopher W. Cianci ,  Moira Hagen ,  Mark Krystal ,  Richard J. Colonno

    IPC分类号： A61K38/00 ,  C07H21/00 ,  C12N15/113 ,  C12N5/00 ,  A61K48/00 ,  C12N15/00 ,  C07N14/00

    CPC分类号： C12N15/1131 ,  C07H21/00 ,  A61K38/00 ,  C12N2310/13 ,  C12N2310/317

    摘要： Novel capped oligonucleotides useful in treatment of influenza infection. A synthetically derived 67-nucleotide RNA substrate, containing a �.sup.32 P! labeled cap-1 structure was used to analyze parameters of influenza virus endonuclease activity. This substrate was specifically cleaved by the influenza virus polymerase to yield a single capped 11-nucleotide fragment capable of directly priming transcription. An analysis of systematic truncations of this RNA substrate in cleavage, elongation, and binding reactions demonstrated that the minimum chain length required for cleavage was one nucleotide past the cleavage site. In contrast, the minimum chain length required for priming activity was found to be 9 nucleotides, while a chain length of at least 4 nucleotides was required for efficient binding. Based on these chain length requirements, the present inventors show that a pool of capped oligonucleotides--too short to prime transcription but long enough to bind with high affinity to the viral polymerase--are potent inhibitors of cap-dependent in vitro transcription.

    摘要翻译： 用于治疗流感感染的新型封端寡核苷酸。 使用含有[32P]标记的cap-1结构的合成衍生的67-核苷酸RNA底物来分析流感病毒核酸内切酶活性的参数。 该底物被流感病毒聚合酶特异地切割，得到能够直接引发转录的单一封端的11-核苷酸片段。 在裂解，延伸和结合反应中对该RNA底物的系统截短的分析表明，切割所需的最短链长度是经过切割位点的一个核苷酸。 相比之下，发现引发活性所需的最短链长度为9个核苷酸，而需要至少4个核苷酸的链长才能有效结合。 基于这些链长度要求，本发明人表明，封闭的寡核苷酸库 - 太短而不能提供转录但足够长以高度结合到病毒聚合酶 - 是帽依赖性体外转录的有效抑制剂。