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公开(公告)号：US09102924B2
公开(公告)日：2015-08-11
申请号：US12425303
申请日：2009-04-16
申请人： Keith A. Bauer , Ellen Fiss , David H. Gelfand , Edward S. Smith , Shawn Suko , Olga Budker , Nancy Schoenbrunner , Susanne Stoffel , Thomas Myers
发明人： Keith A. Bauer , Ellen Fiss , David H. Gelfand , Edward S. Smith , Shawn Suko , Olga Budker , Nancy Schoenbrunner , Susanne Stoffel , Thomas Myers
IPC分类号： C12N9/12
CPC分类号： C12N9/1252 , C12N9/1276 , C12P19/34
摘要： Disclosed are mutant DNA polymerases having improved extension rates relative to a corresponding, unmodified polymerase. The mutant polymerases are useful in a variety of disclosed primer extension methods. The mutant polymerases overcome the inhibitory effects by an intercalating dye. Therefore, the mutant polymerases are useful in a variety of disclosed methods in combination with an intercalating dye. Also disclosed are related compositions, including recombinant nucleic acids, vectors, and host cells, which are useful, e.g., for production of the mutant DNA polymerases.
摘要翻译： 公开了相对于相应的未修饰聚合酶具有改进的延伸速率的突变型DNA聚合酶。 突变型聚合酶可用于各种公开的引物延伸方法。 突变型聚合酶克服了嵌入染料的抑制作用。 因此，突变型聚合酶可与各种公开的方法结合插入染料使用。 还公开了相关组合物，包括重组核酸，载体和宿主细胞，其可用于例如用于产生突变型DNA聚合酶。
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公开(公告)号：US20090280539A1
公开(公告)日：2009-11-12
申请号：US12425303
申请日：2009-04-16
申请人： Keith A. Bauer , Ellen Fiss , David H. Gelfand , Edward S. Smith , Shawn Suko , Olga Budker , Nancy Schoenbrunner , Susanne Stoffel
发明人： Keith A. Bauer , Ellen Fiss , David H. Gelfand , Edward S. Smith , Shawn Suko , Olga Budker , Nancy Schoenbrunner , Susanne Stoffel
CPC分类号： C12N9/1252 , C12N9/1276 , C12P19/34
摘要： Disclosed are mutant DNA polymerases having improved extension rates relative to a corresponding, unmodified polymerase. The mutant polymerases are useful in a variety of disclosed primer extension methods. The mutant polymerases overcome the inhibitory effects by an intercalating dye. Therefore, the mutant polymerases are useful in a variety of disclosed methods in combination with an intercalating dye. Also disclosed are related compositions, including recombinant nucleic acids, vectors, and host cells, which are useful, e.g., for production of the mutant DNA polymerases.
摘要翻译： 公开了相对于相应的未修饰聚合酶具有改进的延伸速率的突变型DNA聚合酶。 突变型聚合酶可用于各种公开的引物延伸方法。 突变型聚合酶克服了嵌入染料的抑制作用。 因此，突变型聚合酶可与各种公开的方法结合插入染料使用。 还公开了相关组合物，包括重组核酸，载体和宿主细胞，其可用于例如用于产生突变型DNA聚合酶。
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3.发明授权Recombinant expression vectors and purification methods for Thermus thermophilus DNA polymerase 失效Thermus thermophilus DNA聚合酶的重组表达载体和纯化方法
公开(公告)号：US5618711A
公开(公告)日：1997-04-08
申请号：US384490
申请日：1995-02-06
CPC分类号： C12N9/1252 , C12N9/1276
摘要： Recombinant DNA sequences encoding the DNA polymerase activity of Thermus thermophilus can be used to construct recombinant vectors and transformed host cells for production of the activity. T. thermophilus DNA polymerase is an .about.94 kDa protein especially useful in the DNA amplification procedure known as the polymerase chain reaction.
摘要翻译： 可以使用编码嗜热栖热菌的DNA聚合酶活性的重组DNA序列构建重组载体和转化的宿主细胞以产生该活性。 嗜热链球菌DNA聚合酶是DIFFERENCE 94 kDa蛋白质，特别适用于称为聚合酶链反应的DNA扩增过程。
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4.发明授权Mutated thermostable nucleic acid polymerase enzyme from thermotoga maritima 失效来自热带海洋假单胞菌的突变的热稳定性核酸聚合酶
公开(公告)号：US5420029A
公开(公告)日：1995-05-30
申请号：US971819
申请日：1993-02-03
IPC分类号： C12N1/21 , C12N9/12 , C12N15/00 , C12N15/09 , C12N15/54 , C12N15/70 , C12P19/34 , C12Q1/68 , C12Q1/70 , C12R1/01 , C12R1/19
CPC分类号： C12N9/1252 , C12N15/70 , C12N9/1276 , C12P19/34 , C12Q1/6876 , C12Q2521/101
摘要： A purified thermostable enzyme is derived from the eubacterium Thermotoga maritima. The enzyme has a molecular weight as determined by gel electrophoresis of about 97 kilodaltons and DNA polymerase I activity. The enzyme can be produced from native or recombinant host cells and can be used with primers and nucleoside triphosphates in a temperature-cycling chain reaction where at least one nucleic acid sequence is amplified in quantity from an existing sequence.
摘要翻译： PCT No.PCT / US91 / 05753 Sec。 371日期：1993年2月3日 102（e）日期1993年2月3日PCT 1991年8月13日PCT PCT。 出版物WO92 / 03556 日期1993年3月5日。纯化的热稳定酶源自海根杆菌。 该酶具有通过约97千道尔顿的凝胶电泳和DNA聚合酶I活性测定的分子量。 酶可以由天然或重组宿主细胞产生，并且可以在温度循环链反应中与引物和核苷三磷酸一起使用，其中至少一个核酸序列的量从现有序列中扩增。
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公开(公告)号：US4889818A
公开(公告)日：1989-12-26
申请号：US063509
申请日：1987-06-17
CPC分类号： C12P19/34 , C12N9/1252 , C12N9/1276
摘要： A purified thermostable enzyme is obtained that has unique characteristics. Preferably the enzyme is isolated from the Thermus aquaticus species and has a molecular weight of about 86,000-90,000 daltons. The thermostable enzyme may be native or recombinant and may be used in a temperature-cycling chain reaction wherein at least one nucleic acid sequence is amplified in quantity from an existing sequence with the aid of selected primers and nucleotide triphosphates. The enzyme is preferably stored in a buffer of non-ionic detergents that lends stability to the enzyme.
摘要翻译： 获得了具有独特特征的纯化的热稳定酶。 优选地，酶从水生栖热菌属物种分离并且具有约86,000-90,000道尔顿的分子量。 热稳定酶可以是天然的或重组的，并且可以用于温度循环链式反应，其中借助于所选择的引物和三磷酸核苷酸，从现有序列中至少一个核酸序列的量被扩增。 该酶优选储存在提供酶稳定性的非离子型洗涤剂的缓冲液中。
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公开(公告)号：US4870013A
公开(公告)日：1989-09-26
申请号：US196789
申请日：1988-05-18
摘要： Expression vectors containing coding sequences under the control of SV40 early and RSV promoters are disclosed as useful in producing proteins in saccharomyces yeasts. Construction of such vectors, and their use in yeast transformations are described.
摘要翻译： 公开了含有SV40早期和RSV启动子控制下的编码序列的表达载体，用于在酵母菌中产生蛋白质。 描述了这些载体的构建及其在酵母转化中的应用。
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7.发明授权Recombinant expression vectors and purification methods for thermus thermophilus DNA polymerase 失效热嗜热杆菌DNA聚合酶的重组表达载体和纯化方法
公开(公告)号：US5789224A
公开(公告)日：1998-08-04
申请号：US459383
申请日：1995-06-02
IPC分类号： C12N9/12
CPC分类号： C12N9/1252 , C12N9/1276
摘要： Recombinant DNA sequences encoding the DNA polymerase activity of Thermus thermophilus can be used to construct recombinant vectors and transformed host cells for production of the activity. T. thermophilus DNA polymerase is an .about.94 kDa protein especially useful in the DNA amplification procedure known as the polymerase chain reaction.
摘要翻译： 可以使用编码嗜热栖热菌的DNA聚合酶活性的重组DNA序列构建重组载体和转化的宿主细胞以产生该活性。 嗜热链球菌DNA聚合酶是DIFFERENCE 94 kDa蛋白质，特别适用于称为聚合酶链反应的DNA扩增过程。
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公开(公告)号：US5352600A
公开(公告)日：1994-10-04
申请号：US971798
申请日：1992-11-05
申请人： David H. Gelfand , Susanne Stoffel
发明人： David H. Gelfand , Susanne Stoffel
CPC分类号： C12P19/34 , C12N15/70 , C12N9/1252 , C12N9/1276
摘要： A purified thermostable enzyme is obtained that has unique characteristics. Preferably the enzyme is isolated from the Thermus aquaticus species and has a molecular weight of about 86,000-95,000 daltons. The thermostable enzyme may be native or recombinant and may be used in a temperature-cycling chain reaction wherein at least one nucleic acid sequence is amplified in quantity from an existing sequence with the aid of selected primers and nucleotide triphosphates. The enzyme is preferably stored in a buffer containing non-ionic detergents that lends stability to the enzyme.
摘要翻译： 获得了具有独特特征的纯化的热稳定酶。 优选地，酶从水生栖热菌属物种分离并且具有约86,000-95,000道尔顿的分子量。 热稳定酶可以是天然的或重组的，并且可以用于温度循环链式反应，其中借助于所选择的引物和三磷酸核苷酸，从现有序列中至少一个核酸序列的量被扩增。 该酶优选储存在含有赋予酶稳定性的非离子型洗涤剂的缓冲液中。
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9.发明授权Selectable fusion protein having aminoglycoside phosphotransferase activity 失效具有氨基糖苷磷酸转移酶活性的可选择的融合蛋白
公开(公告)号：US5116750A
公开(公告)日：1992-05-26
申请号：US223430
申请日：1988-07-22
CPC分类号： C12N15/63 , C07K14/55 , C12N15/70 , C12N9/12 , C07K2319/00 , C07K2319/61 , Y10S930/31
摘要： A novel a selectable fusion protein having aminoglycoside phosphotransferase activity is disclosed. The marker comprises the coding sequences for aminoglycoside phosphotransferase I (APH-I) which has been modified and truncated so as to render its use in recombinant vectors more convenient. The modified, truncated sequence (mtAPH-I) gene is capable, upon expression, of conferring resistance to a number of antibiotics on the host. One of these antibiotics, G418, is toxic to eucaryotic as well as procaryotic hosts. Also disclosed are methods of constructing fusion proteins having N-terminal sequences corresponding to a desired peptide sequence, and C-terminal sequences comprising the amino acids encoded by mtAPH-I. The preferred N-terminal sequences are the first 11 amino acids of .beta.-isopropyl malate dehydrogenase, and the first 7 amino acids of yeast enolase.
摘要翻译： 公开了一种具有氨基糖苷磷酸转移酶活性的新型可选择融合蛋白。 该标记包括已被修饰和截短的氨基糖苷磷酸转移酶I（APH-1）的编码序列，使其在重组载体中的使用更加方便。 修饰的截短序列（mtAPH-I）基因在表达时能够赋予宿主对许多抗生素的抗性。 其中一种抗生素G418对真核生物以及原核生物有毒性。 还公开了构建具有对应于所需肽序列的N末端序列的融合蛋白的方法，以及包含由mtAPH-I编码的氨基酸的C-末端序列。 优选的N-末端序列是苹果酸β-异丙酯脱氢酶的前11个氨基酸，酵母烯醇化酶的前7个氨基酸。
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10.发明授权Stabilized thermostable nucleic acid polymerase compositions containing non-ionic polymeric detergents 失效含有非离子聚合物洗涤剂的稳定的耐热核酸聚合酶组合物
公开(公告)号：US6127155A
公开(公告)日：2000-10-03
申请号：US873897
申请日：1992-04-24
CPC分类号： C12P19/34 , C12N15/70 , C12N9/1252 , C12Q1/686 , Y10S435/814
摘要： A purified thermostable nucleic acid polymerase is obtained that has unique characteristics. Preferably the nucleic acid polymerase is DNA polymerase isolated from a Thermus aquaticus species and has a molecular weight of about 86,000-95,000 daltons. The thermostable nucleic acid polymerase may be native or recombinant and may be used in a temperature-cycling chain reaction wherein at least one nucleic acid sequence is amplified in quantity from an existing sequence with the aid of selected primers and nucleotide triphosphates. The nucleic acid polymerase is preferably stored in a buffer containing non-ionic detergents that lends stability to the nucleic acid polymerase. A preferred buffer contains glycerol, polyoxyethylated sorbitan monolaurate, ethoxylated nonyl phenol and gelatin.
摘要翻译： 获得了具有独特特征的纯化的热稳定性核酸聚合酶。 优选地，所述核酸聚合酶是从栖热栖热菌属物种分离的DNA聚合酶，其分子量约为86,000-95,000道尔顿。 热稳定性核酸聚合酶可以是天然的或重组的，并且可以用于温度循环链式反应，其中借助于选定的引物和三磷酸核苷酸，从现存的序列中扩增至少一个核酸序列。 核酸聚合酶优选储存在含有使核酸聚合酶具有稳定性的非离子型清净剂的缓冲液中。 优选的缓冲液含有甘油，聚氧乙基化脱水山梨糖醇单月桂酸酯，乙氧基化壬基苯酚和明胶。
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