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21. 发明授权

US06252136B1 Transgenic organisms having tetracycline-regulated transcriptional regulatory systems 有权
标题翻译：具有四环素调节转录调控系统的转基因生物
公开(公告)号：US06252136B1
公开(公告)日：2001-06-26
申请号：US09163269
申请日：1998-09-29
申请人： Hermann Bujard , Manfred Gossen , Jochen G. Salfeld , Jeffrey W. Voss
发明人： Hermann Bujard , Manfred Gossen , Jochen G. Salfeld , Jeffrey W. Voss
IPC分类号： C12N1509
CPC分类号： C12N15/62 , A01K67/0275 , A01K2217/05 , A01K2217/075 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N15/635 , C12N15/85 , C12N2830/006
摘要： Transgenic animals carrying two transgenes, the first coding for a transactivator fusion protein comprising a tet repressor and a polypeptide which directly or indirectly activates in eucaryotic cells, and the second comprising a gene operably linked to a minimal promoter operably linked to at least one tet operator sequence, are disclosed. Isolated DNA molecules (e.g., targeting vectors) for integrating a polynucleotide sequence encoding a transactivator of the invention at a predetermined location within a second target DNA molecule by homologous recombination are also disclosed. Transgenic animals having the DNA molecules of the invention integrated at a predetermined location in a chromosome by homologous recombination are also encompassed by the invention. Methods to regulate the expression of a tet operator linked-gene of interest by administering tetracycline or a tetracycline analogue to an animal of the invention are also disclosed. The regulatory system of the invention allows for conditional inactivation or modulation of expression of a gene of interest in a host cell or animal.
摘要翻译：携带两个转基因的转基因动物，第一编码包含直接或间接在真核细胞中活化的tet阻遏物和多肽的反式激活因子融合蛋白，第二编码包含与可操作地连接至少一个tet操纵子的最小启动子可操作地连接的基因序列。还公开了通过同源重组将编码本发明的反式激活物的多核苷酸序列整合到第二靶DNA分子内的预定位置的分离的DNA分子（例如靶向载体）。通过同源重组将具有整合在染色体中预定位置的本发明的DNA分子的转基因动物也包括在本发明中。还公开了通过向本发明的动物施用四环素或四环素类似物来调节所述目的基因的表达的方法。本发明的调节系统允许在宿主细胞或动物中有条件地失活或调控感兴趣的基因的表达。

22. 发明授权

US5912411A Mice transgenic for a tetracycline-inducible transcriptional activator 失效
标题翻译：四环素诱导型转录激活剂转基因小鼠
公开(公告)号：US5912411A
公开(公告)日：1999-06-15
申请号：US487472
申请日：1995-06-07
申请人： Hermann Bujard , Manfred Gossen
发明人： Hermann Bujard , Manfred Gossen
IPC分类号： A01K67/027 , A61K48/00 , C07K14/00 , C07K14/035 , C07K14/195 , C07K14/245 , C07K19/00 , C12N15/11 , C12N15/62 , C12N15/63 , C12N15/67 , C12N15/85 , C12N5/00 , C12N15/00 , C12N15/09
CPC分类号： C07K14/005 , A01K67/0275 , C07K14/245 , C12N15/62 , C12N15/63 , C12N15/635 , C12N15/67 , C12N15/85 , C12N15/8509 , A01K2217/05 , A01K2217/075 , A01K2217/20 , A01K2227/105 , A01K2267/01 , A01K2267/02 , A01K2267/03 , A01K2267/0393 , A61K48/00 , C07K2319/00 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N2710/16622 , C12N2800/30 , C12N2830/00 , C12N2830/003 , C12N2830/006 , C12N2830/32
摘要： Transgenic mice carrying a transgene comprising a nucleic acid molecule encoding protein useful for regulating the expression of genes in eukaryotic cells in a highly controlled manner are disclosed. In the regulatory system of the invention, transcription of a tet operator-linked nucleotide sequence is stimulated by a transcriptional activator fusion protein composed of two polypeptides, a first polypeptide which binds to tet operator sequences in the presence of tetracycline operatively linked to a second polypeptide activates transcription in eukaryotic cells. In a preferred embodiment, the transgene encoding the transcriptional activator fusion protein is integrated at a predetermined location within the chromosome of the transgenic mouse.
摘要翻译：公开了携带转基因的转基因，该转基因包含可用于以高度受控的方式调控真核细胞中的基因表达的编码蛋白的核酸分子。在本发明的调节系统中，tet操纵子连接的核苷酸序列的转录受到由两个多肽组成的转录激活物融合蛋白的刺激，所述两个多肽是在与第二个多肽有效连接的四环素存在下结合tet操纵子序列的第一多肽在真核细胞中激活转录。在优选的实施方案中，编码转录激活物融合蛋白的转基因整合在转基因小鼠的染色体内的预定位置。

23. 发明授权

US5814618A Methods for regulating gene expression 失效
标题翻译：调节基因表达的方法
公开(公告)号：US5814618A
公开(公告)日：1998-09-29
申请号：US485978
申请日：1995-06-07
申请人： Hermann Bujard , Manfred Gossen
发明人： Hermann Bujard , Manfred Gossen
IPC分类号： A01K67/027 , A61K48/00 , C07K14/00 , C07K14/035 , C07K14/195 , C07K14/245 , C07K19/00 , C12N15/11 , C12N15/63 , C12N15/67 , C12N15/85 , C12N15/00
CPC分类号： C07K14/005 , A01K67/0275 , C07K14/245 , C12N15/62 , C12N15/63 , C12N15/635 , C12N15/67 , C12N15/85 , C12N15/8509 , A01K2217/05 , A01K2217/075 , A01K2217/20 , A01K2227/105 , A01K2267/0393 , A61K48/00 , C07K2319/00 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N2710/16622 , C12N2830/003 , C12N2830/006 , C12N2830/32
摘要： Methods of regulating gene expression in subjects using tetracycline-responsive fusion proteins are disclosed. In one embodiment, the method involves introducing into a cell the subject a nucleic acid molecule encoding a fusion protein which inhibits transcription, the fusion protein comprising a first polypeptide which binds to a tet operator sequence, operatively linked to a heterologous second polypeptide which inhibits transcription in eukaryotic cells; and modulating the concentration of a tetracycline, or analogue thereof, in the subject. The first polypeptide can binds to a tet operator sequence in the absence, but not the presence, of tetracycline. Alternatively, the first polypeptide can binds to a tet operator sequence in the presence, but not the absence, of tetracycline. In another embodiment, the method of the invention involves obtaining a cell from a subject, introducing into the cell a first nucleic acid molecule which operatively links a gene to at least one tet operator sequence, introducing into the cell a second nucleic acid molecule encoding an inhibitory fusion protein of the invention to form a modified cell, administering the modified cell to the subject and modulating the concentration of a tetracycline, or analogue thereof, in the subject. The first and second nucleic acid molecules can be linked or can be separate molecules.
摘要翻译：公开了使用四环素响应融合蛋白调节受试者基因表达的方法。在一个实施方案中，该方法包括向受试者引入编码抑制转录的融合蛋白的核酸分子，该融合蛋白包含与tet操纵子序列结合的第一多肽，该第一多肽与抑制转录的异源第二多肽有效连接在真核细胞中并调节受试者中四环素或其类似物的浓度。在不存在但不存在四环素的情况下，第一种多肽可以结合tet操纵子序列。或者，第一多肽可以在四环素存在下但不存在时与tet操纵子序列结合。在另一个实施方案中，本发明的方法涉及从受试者获得细胞，将在第一核酸分子中引入至少一个tet操纵子序列的第一核酸分子，将编码本发明的抑制性融合蛋白以形成修饰的细胞，向受试者施用修饰的细胞并调节受试者中四环素或其类似物的浓度。第一和第二核酸分子可以连接或可以是分开的分子。

24. 发明授权

US5650298A Tight control of gene expression in eucaryotic cells by tetracycline-responsive promoters 失效
标题翻译：通过四环素响应启动子对真核细胞中基因表达的严格控制
公开(公告)号：US5650298A
公开(公告)日：1997-07-22
申请号：US260452
申请日：1994-06-14
申请人： Hermann Bujard , Manfred Gossen , Jochen G. Salfeld , Jeffrey W. Voss
发明人： Hermann Bujard , Manfred Gossen , Jochen G. Salfeld , Jeffrey W. Voss
IPC分类号： A01K67/027 , A61K48/00 , C07K14/00 , C12N5/08 , C12N5/10 , C12N15/00 , C12N15/09 , C12N15/31 , C12N15/62 , C12N15/63 , C12N15/85 , C12R1/91 , C12P21/00 , C07H21/04
CPC分类号： A01K67/0275 , C12N15/62 , C12N15/635 , C12N15/85 , A01K2217/05 , A01K2217/075 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N2830/006
摘要： Transgenic animals carrying two transgenes, the first coding for a transactivator fusion protein comprising a tet repressor and a polypeptide which directly or indirectly activates in eucaryotic cells, and the second comprising a gene operably linked to a minimal promoter operably linked to at least one tet operator sequence, are disclosed. Isolated DNA molecules (e.g., targeting vectors) for integrating a polynucleotide sequence encoding a transactivator of the invention at a predetermined location within a second target DNA molecule by homologous recombination are also disclosed. Transgenic animals having the DNA molecules of the invention integrated at a predetermined location in a chromosome by homologous recombination are also encompassed by the invention. Methods to regulate the expression of a tet operator linked-gene of interest by administering tetracycline or a tetracycline analogue to an animal of the invention are also disclosed. The regulatory system of the invention allows for conditional inactivation or modulation of expression of a gene of interest in a host cell or animal.
摘要翻译：携带两个转基因的转基因动物，第一编码包含直接或间接在真核细胞中活化的tet阻遏物和多肽的反式激活因子融合蛋白，第二编码包含与可操作地连接至少一个tet操纵子的最小启动子可操作地连接的基因序列。还公开了通过同源重组将编码本发明的反式激活物的多核苷酸序列整合到第二靶DNA分子内的预定位置的分离的DNA分子（例如靶向载体）。通过同源重组将具有整合在染色体中预定位置的本发明的DNA分子的转基因动物也包括在本发明中。还公开了通过向本发明的动物施用四环素或四环素类似物来调节所述目的基因的表达的方法。本发明的调节系统允许在宿主细胞或动物中有条件地失活或调控感兴趣的基因的表达。

25. 发明授权

US5589362A Tetracycline regulated transcriptional modulators with altered DNA binding specificities 失效
标题翻译：具有改变的DNA结合特异性的四环素调节的转录调节子
公开(公告)号：US5589362A
公开(公告)日：1996-12-31
申请号：US485971
申请日：1995-06-07
申请人： Hermann Bujard , Manfred Gossen , Wolfgang Hillen , Vera Helbl , Dirk Schnappinger
发明人： Hermann Bujard , Manfred Gossen , Wolfgang Hillen , Vera Helbl , Dirk Schnappinger
IPC分类号： A01K67/027 , A61K48/00 , C07K14/00 , C07K14/035 , C07K14/195 , C07K14/245 , C07K19/00 , C12N15/11 , C12N15/63 , C12N15/67 , C12N15/85 , C12P21/00 , C07H21/04 , C12N15/31
CPC分类号： A01K67/0275 , C07K14/005 , C07K14/245 , C12N15/63 , C12N15/635 , C12N15/67 , C12N15/85 , C12N15/8509 , A01K2217/05 , A01K2217/075 , A01K2217/20 , A01K2227/105 , A01K2267/0393 , A61K48/00 , C07K2319/00 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N2710/16622 , C12N2830/003 , C12N2830/006 , C12N2830/30 , C12N2830/32
摘要： Isolated nucleic acid molecules encoding fusion proteins which regulate transcription in eukaryotic cells are disclosed. The fusion proteins of the invention comprise a Tet repressor having at least one amino acid mutation that confers on the fusion protein an ability to bind a class B tet operator sequence having a nucleotide substitution at position +4 or +6, operatively linked to a polypeptide which regulates transcription in eukaryotic cells. Methods for regulating transcription of a tet operator-linked gene in a cell are also provided. In one embodiment, the method involves introducing into the cell a nucleic acid molecule encoding a fusion protein which regulates transcription, the fusion protein comprising a Tet repressor having at least one amino acid mutation that confers on the fusion protein an ability to bind a class B tet operator sequence having a nucleotide substitution at position +4 or +6, operatively linked to a polypeptide which regulates transcription in eukaryotic cells, and modulating the concentration of a tetracycline, or analogue thereof, in contact with the cell.
摘要翻译：公开了编码在真核细胞中调节转录的融合蛋白的分离的核酸分子。本发明的融合蛋白包含Tet阻遏物，其具有赋予融合蛋白的至少一个氨基酸突变，其结合具有与多肽有效连接的位置+4或+6的核苷酸取代的B类tet操纵子序列的能力其调控真核细胞中的转录。还提供了调节细胞中tet操纵子连接基因转录的方法。在一个实施方案中，该方法包括将编码调节转录的融合蛋白的核酸分子导入细胞，所述融合蛋白包含具有至少一个氨基酸突变的Tet阻遏物，赋予融合蛋白具有结合B类的能力 tet操纵子序列在+4或+6位具有核苷酸取代，可操作地连接到调节真核细胞转录的多肽，并调节与细胞接触的四环素或其类似物的浓度。

26. 发明申请

US20060127413A1 Recombinant mva strains as potential vaccines against p. falciparum malaria 审中-公开
标题翻译：重组mva菌株作为针对p的潜在疫苗。恶性疟疾
公开(公告)号：US20060127413A1
公开(公告)日：2006-06-15
申请号：US10532067
申请日：2003-09-26
申请人： Gerd Sutter , Hermann Bujard , Nicole Westerfeld , Miao Jun
发明人： Gerd Sutter , Hermann Bujard , Nicole Westerfeld , Miao Jun
IPC分类号： A61K39/12 , C12N7/00 , C12N15/86
CPC分类号： C12N15/86 , A61K39/015 , A61K2039/5256 , C12N2710/24143 , Y02A50/412
摘要： This invention relates to recombinant viruses based on MVA, which comprise at least one nucleic acid coding for a Plasmodium falciparum MSP-1 protein, a fragment or mutein of it. Furthermore, methods for the production of the recombinant viruses, virus-containing vaccines and the use of the recombinant viruses for the prophylaxis and/or therapy of malaria are provided.
摘要翻译：本发明涉及基于MVA的重组病毒，其包含至少一种编码恶性疟原虫MSP-1蛋白质的核酸，其片段或突变蛋白。此外，提供了用于生产重组病毒，含病毒疫苗和使用重组病毒用于预防和/或治疗疟疾的方法。

27. 发明授权

US06914124B2 Tetracycline-regulated transcriptional activator fusion proteins 失效
标题翻译：四环素调节的转录激活融合蛋白
公开(公告)号：US06914124B2
公开(公告)日：2005-07-05
申请号：US09777317
申请日：2001-02-05
申请人： Hermann Bujard , Manfred Gossen
发明人： Hermann Bujard , Manfred Gossen
IPC分类号： A01K67/027 , A61K48/00 , C07K14/00 , C12N5/08 , C12N5/10 , C12N15/00 , C12N15/09 , C12N15/31 , C12N15/62 , C12N15/63 , C12N15/85 , C12R1/91
CPC分类号： A01K67/0275 , A01K2217/05 , A01K2217/075 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N15/62 , C12N15/635 , C12N15/85 , C12N2830/006
摘要： A method for regulating expression of a tet operator-linked gene in a cell of a subject is disclosed. In one embodiment, the method involves introducing into the cell a nucleic acid molecule encoding a tetracycline-controllable transactivator (tTA), the tTA comprising a Tet repressor operably linked to a polypeptide which directly or indirectly activates transcription in eucaryotic cells; and modulating the concentration of a tetracycline, or analogue thereof, in the subject. Alternatively, in another embodiment, the method involves obtaining the cell from the subject, introducing into the cell a first nucleic acid molecule which operatively links a gene to at least one tet operator sequence, introducing into the cell a second nucleic acid molecule encoding a tTA, to form a modified cell, administering the modified cell to the subject, and modulating the concentration of a tetracycline, or analogue thereof, in the subject. The first and second nucleic acid molecule can be within a single molecule (e.g., in the same vector) or on separate molecules.
摘要翻译：公开了一种用于调节受试者细胞中tet操纵子连接基因表达的方法。在一个实施方案中，所述方法包括将编码四环素可控反式激活因子（tTA）的核酸分子引入细胞，所述tTA包含可操作地连接到直接或间接激活真核细胞中的转录的多肽的Tet阻遏物; 并调节受试者中四环素或其类似物的浓度。或者，在另一个实施方案中，所述方法包括从受试者获得细胞，向细胞中引入第一核酸分子，该第一核酸分子可操作地将基因连接至至少一个tet操纵子序列，将编码tTA的第二核酸分子，以形成修饰的细胞，向受试者施用修饰的细胞，并调节受试者中四环素或其类似物的浓度。第一和第二核酸分子可以在单分子内（例如，在同一载体中）或分开的分子上。

28. 发明申请

US20050095256A1 Recombinant process for preparing a complete malaria antigen gp190/MSP1 审中-公开
标题翻译：用于制备完整的疟疾抗原gp190 / MSP1的重组方法
公开(公告)号：US20050095256A1
公开(公告)日：2005-05-05
申请号：US10954924
申请日：2004-09-29
申请人： Hermann Bujard , Ralf Tolle , Weiqing Pan
发明人： Hermann Bujard , Ralf Tolle , Weiqing Pan
IPC分类号： C12N15/09 , A61K39/00 , A61K39/15 , A61P33/06 , C07K14/445 , C12N1/10 , C12N1/11 , C12N1/19 , C12N1/21 , C12N5/10 , C12N15/02 , C12N15/30 , C12P21/08 , C12R1/19 , A61K39/002 , C07H21/04 , C12N15/74
CPC分类号： C07K14/445 , A61K39/00 , A61K2039/51 , A61K2039/523 , A61K2039/53 , C07K2319/00 , Y02A50/41 , Y02A50/412
摘要： Recombinant manufacturing process for a complete malaria antigen gp190/MSP1 The invention concerns a recombinant manufacturing process for the complete gp190/MSP1 surface protein of Plasmodium, in particular P. falciparum, as well as the complete DNA sequence of this protein and suitable host organisms for expression of the sequence, by means of which the protein in its entirety can for the first time be synthesized outside the parasite. The invention opens up for the first time the possibility of manufacturing the gp190/MSP1 surface protein in sufficient quantity; in addition there is the matter of making gp190/MSP1 available as a vaccine. Finally the invention sets out a process for the stabilization of AT-rich genes (FIG. 1).
摘要翻译：用于完整疟疾抗原gp190 / MSP1的重组制备方法本发明涉及用于疟原虫，特别是恶性疟原虫的完整gp190 / MSP1表面蛋白的重组制备方法，以及该蛋白质的完整DNA序列和适合的宿主生物体蛋白质整体可以首次在寄生虫外合成序列的表达。本发明首次开放了足够数量制造gp190 / MSP1表面蛋白的可能性; 另外还有可能使gp190 / MSP1作为疫苗使用。最后，本发明提出了一种稳定AT富含基因的方法（图1）。

29. 发明授权

US06242667B1 Transgenic organisms having tetracycline-regulated transcriptional regulatory systems 有权
标题翻译：具有四环素调节转录调控系统的转基因生物
公开(公告)号：US06242667B1
公开(公告)日：2001-06-05
申请号：US09161902
申请日：1998-09-28
申请人： Hermann Bujard , Manfred Gossen
发明人： Hermann Bujard , Manfred Gossen
IPC分类号： C12N1509
CPC分类号： C07K14/005 , A01K67/0275 , A01K2217/05 , A01K2217/075 , A01K2217/20 , A01K2227/105 , A01K2267/01 , A01K2267/02 , A01K2267/03 , A01K2267/0393 , A61K48/00 , C07K14/245 , C07K2319/00 , C07K2319/09 , C07K2319/61 , C07K2319/71 , C07K2319/80 , C12N15/85 , C12N15/8509 , C12N2710/16622 , C12N2800/30 , C12N2830/00 , C12N2830/003 , C12N2830/006 , C12N2830/32
摘要： Transgenic animals carrying a transgene comprising a nucleic acid molecule encoding protein useful for regulating the expression of genes in eukaryotic cells and organisms in a highly controlled manner are disclosed. In the regulatory system of the invention, transcription of a tet operator-linked nucleotide sequence is stimulated by a transcriptional activator fusion protein composed of two polypeptides, a first polypeptide which binds to tet operator sequences in the presence of tetracycline operatively linked to a second polypeptide activates transcription in eukaryotic cells. In a preferred embodiment, the transgene encoding the transcriptional activator fusion protein is integrated at a predetermined location within the chromosome of the transgenic animal.
摘要翻译：公开了携带转基因的转基因，其包含编码可用于以高度受控的方式调控真核细胞和生物体中的基因表达的蛋白质的核酸分子。在本发明的调节系统中，tet操纵子连接的核苷酸序列的转录受到由两个多肽组成的转录激活物融合蛋白的刺激，所述两个多肽是在与第二个多肽有效连接的四环素存在下结合tet操纵子序列的第一多肽在真核细胞中激活转录。在优选的实施方案中，编码转录激活物融合蛋白的转基因整合在转基因动物的染色体内的预定位置。

30. 发明授权

US6087166A Transcriptional activators with graded transactivation potential 失效
标题翻译：具有分级反式激活潜能的转录激活子
公开(公告)号：US6087166A
公开(公告)日：2000-07-11
申请号：US888080
申请日：1997-07-03
申请人： Udo Baron , Manfred Gossen , Hermann Bujard
发明人： Udo Baron , Manfred Gossen , Hermann Bujard
IPC分类号： A01K67/027 , A61K48/00 , C07K14/035 , C07K14/72 , C07K19/00 , C12N1/15 , C12N1/19 , C12N1/21 , C12N5/10 , C12N15/09 , C12N15/63 , C12N15/85 , C12N1/00 , C12N15/62
CPC分类号： C07K14/005 , C12N15/635 , C12N15/85 , A01K2217/05 , A61K48/00 , C07K2319/00 , C12N2710/16622 , C12N2830/003 , C12N2830/006
摘要： Transcriptional activators which differ in their activation potential by more than 3 orders of magnitude are provided. The transactivators are fusions between a DNA binding protein (e.g., a Tet repressor) and minimal transcriptional activation domains derived from Herpes simplex virus protein 16 (VP16). Substitution mutations at amino acid position 442 within the minimal VP16 domain provide transactivators with differing transactivation ability. Moreover, chimeric activation domains comprising both wild type and mutant minimal VP16 domains provide additional variants with differing transactivation ability. Various aspects of the invention pertain to nucleic acid molecules, vectors, host cells, fusion proteins, transgenic and homologous recombinant organisms and methods of regulating gene transcription.
摘要翻译：提供了其激活电位不同于3个数量级的转录激活子。反式激活因子是DNA结合蛋白（例如Tet阻遏物）和衍生自单纯疱疹病毒蛋白16（VP16）的最小转录激活结构域之间的融合物。在最小VP16结构域内的氨基酸位置442处的取代突变体提供具有不同反式激活能力的反式激活因子。此外，包含野生型和突变型最小VP16结构域的嵌合活化域提供了具有不同反式激活能力的另外的变体。本发明的各个方面涉及核酸分子，载体，宿主细胞，融合蛋白，转基因和同源重组生物以及调节基因转录的方法。

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