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1. 发明申请

US20110046201A1 METHODS AND COMPOSITIONS FOR SEAMLESS CLONING OF NUCLEIC ACID MOLECULES 审中-公开
标题翻译：无酸克隆核酸分子的方法和组合物
公开(公告)号：US20110046201A1
公开(公告)日：2011-02-24
申请号：US12112803
申请日：2008-04-30
申请人： Jonathan D. Chesnut , Knut R. Madden , Miroslav Dudas , Louis Leong , Adam N. Harris
发明人： Jonathan D. Chesnut , Knut R. Madden , Miroslav Dudas , Louis Leong , Adam N. Harris
IPC分类号： A61K31/713 , C12P19/34 , C12N5/10 , C12N15/63
CPC分类号： C12P19/34 , C07H21/04 , C12N9/90 , C12N15/10 , C12N15/111 , C12N15/64 , C12N15/66 , C12N15/86 , C12N2310/111 , C12N2310/14 , C12N2330/30 , C12N2740/16043 , C12N2800/30 , C12N2800/70 , C12Y599/01
摘要： The present invention is in the fields of biotechnology and molecular biology. More particularly, the present invention relates to cloning or subcloning one or more nucleic acid molecules comprising one or more type IIs restriction enzyme recognition sites. The present invention also embodies cloning such nucleic acid molecules using recombinational cloning methods such as those employing recombination sites and recombination proteins. The present invention also relates to nucleic acid molecules (including RNA and iRNA), as well as proteins, expressed from host cells produced using the methods of the present invention.
摘要翻译：本发明涉及生物技术和分子生物学领域。更具体地说，本发明涉及克隆或亚克隆一个或多个包含一个或多个II型限制酶识别位点的核酸分子。本发明还使用重组克隆方法（例如使用重组位点和重组蛋白的那些）来体现克隆这种核酸分子。本发明还涉及由使用本发明的方法生产的宿主细胞表达的核酸分子（包括RNA和iRNA）以及蛋白质。

2. 发明申请

US20120149069A1 METHODS AND COMPOSITIONS FOR SEAMLESS CLONING OF NUCLEIC ACID MOLECULES 有权
标题翻译：无酸克隆核酸分子的方法和组合物
公开(公告)号：US20120149069A1
公开(公告)日：2012-06-14
申请号：US13401680
申请日：2012-02-21
申请人： Jonathan D. Chesnut , Knut R. Madden , Miroslav Dudas , Louis Leong , Adam N. Harris
发明人： Jonathan D. Chesnut , Knut R. Madden , Miroslav Dudas , Louis Leong , Adam N. Harris
IPC分类号： C12N15/66
CPC分类号： C12P19/34 , C07H21/04 , C12N9/90 , C12N15/10 , C12N15/111 , C12N15/64 , C12N15/66 , C12N15/86 , C12N2310/111 , C12N2310/14 , C12N2330/30 , C12N2740/16043 , C12N2800/30 , C12N2800/70 , C12Y599/01
摘要： The present invention is in the fields of biotechnology and molecular biology. More particularly, the present invention relates to cloning or subcloning one or more nucleic acid molecules comprising one or more type IIs restriction enzyme recognition sites. The present invention also embodies cloning such nucleic acid molecules using recombinational cloning methods such as those employing recombination sites and recombination proteins. The present invention also relates to nucleic acid molecules (including RNA and iRNA), as well as proteins, expressed from host cells produced using the methods of the present invention.
摘要翻译：本发明涉及生物技术和分子生物学领域。更具体地说，本发明涉及克隆或亚克隆一个或多个包含一个或多个II型限制酶识别位点的核酸分子。本发明还使用重组克隆方法（例如使用重组位点和重组蛋白的那些）来体现克隆这种核酸分子。本发明还涉及由使用本发明的方法生产的宿主细胞表达的核酸分子（包括RNA和iRNA）以及蛋白质。

3. 发明授权

US06916632B2 Methods and reagents for molecular cloning 有权
标题翻译：分子克隆的方法和试剂
公开(公告)号：US06916632B2
公开(公告)日：2005-07-12
申请号：US09935280
申请日：2001-08-21
申请人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
发明人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
IPC分类号： A01K67/027 , C12N1/21 , C12N5/10 , C12N15/09 , C12N15/10 , C12N15/64 , C12N15/66 , C40B40/02 , C40B50/06 , C12P19/34 , C12N9/90 , C12N15/00 , C12P21/03
CPC分类号： C12N15/66 , C12N15/10 , C12N15/64
摘要： The present invention provides compositions, methods, and kits for covalently linking nucleic acid molecules. The methods include a strand invasion step, and the compositions and kits are useful for performing such methods. For example, a method of covalently linking double stranded (ds) nucleic acid molecules can include contacting a first ds nucleic acid molecule, which has a topoisomerase linked to a 3′ terminus of one end and has a single stranded 5′ overhang at the same end, with a second ds nucleic acid molecule having a blunt end, such that the 5′ overhang can hybridize to a complementary sequence of the blunt end of the second nucleic acid molecule, and the topoisomerase can covalently link the ds nucleic acid molecules. The methods are simpler and more efficient than previous methods for covalently linking nucleic acid sequences, and the compositions and kits facilitate practicing the methods, including methods of directionally linking two or more ds nucleic acid molecules.
摘要翻译：本发明提供用于共价连接核酸分子的组合物，方法和试剂盒。所述方法包括线入侵步骤，组合物和试剂盒对于进行这些方法是有用的。例如，共价连接双链（ds）核酸分子的方法可以包括使第一个ds核酸分子接触，该第一个ds核酸分子具有与一个末端的3'末端连接的拓扑异构酶并且具有相同的单链（5'）突出端末端，具有平末端的第二ds核酸分子，使得5'突出端可以与第二核酸分子的平端的互补序列杂交，并且拓扑异构酶可以共价连接ds核酸分子。该方法比先前用于共价连接核酸序列的方法更简单和更有效，并且组合物和试剂盒有助于实施该方法，包括定向连接两个或多个ds核酸分子的方法。

4. 发明授权

US6017754A System for isolating and identifying eukaryotic cells transfected with genes and vectors, host cells and methods thereof 失效
标题翻译：用于分离和鉴定用基因和载体转染的真核细胞的系统，宿主细胞及其方法
公开(公告)号：US6017754A
公开(公告)日：2000-01-25
申请号：US518835
申请日：1995-08-24
申请人： Jonathan D. Chesnut , James P. Hoeffler
发明人： Jonathan D. Chesnut , James P. Hoeffler
IPC分类号： C07K16/00 , C12N15/64 , C12N15/65 , C12Q1/68 , C12N5/10 , G01N33/566
CPC分类号： C07K16/00 , C12N15/64 , C12N15/65 , Y10S435/975
摘要： The present invention relates a novel expression system which allows the study of experimental genes of interest on cellular events soon after transfection. The expression system includes a vector which encodes for a recombinant antibody binding unit (rAb). The expression system enables identification and selection of transfected cells from culture to be carried out immediately, within hours, after the transfection
摘要翻译：本发明涉及一种新的表达系统，其允许在转染后不久研究感兴趣的实验基因的细胞事件。表达系统包括编码重组抗体结合单元（rAb）的载体。表达系统使得能够在转染后数小时内立即进行来自培养物的转染细胞的鉴定和选择

5. 发明申请

US20090328244A1 METHODS AND REAGENTS FOR MOLECULAR CLONING 审中-公开
标题翻译：分子克隆的方法和试剂
公开(公告)号：US20090328244A1
公开(公告)日：2009-12-31
申请号：US12409326
申请日：2009-03-23
申请人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
发明人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
IPC分类号： A01K67/027 , C12P19/34 , C12N5/06 , C12N15/11
CPC分类号： C12N15/66 , C12N15/10 , C12N15/64
摘要： The present invention provides compositions, methods, and kits for covalently linking nucleic acid molecules. The methods include a strand invasion step, and the compositions and kits are useful for performing such methods. For example, a method of covalently linking double stranded (ds) nucleic acid molecules can include contacting a first ds nucleic acid molecule, which has a topoisomerase linked to a 3′ terminus of one end and has a single stranded 5′ overhang at the same end, with a second ds nucleic acid molecule having a blunt end, such that the 5′ overhang can hybridize to a complementary sequence of the blunt end of the second nucleic acid molecule, and the topoisomerase can covalently link the ds nucleic acid molecules. The methods are simpler and more efficient than previous methods for covalently linking nucleic acid sequences, and the compositions and kits facilitate practicing the methods, including methods of directionally linking two or more ds nucleic acid molecules.
摘要翻译：本发明提供用于共价连接核酸分子的组合物，方法和试剂盒。所述方法包括线入侵步骤，组合物和试剂盒对于进行这些方法是有用的。例如，共价连接双链（ds）核酸分子的方法可以包括使第一个ds核酸分子接触，该第一个ds核酸分子具有与一个末端的3'末端连接的拓扑异构酶并且具有相同的单链（5'）突出端末端，具有平末端的第二ds核酸分子，使得5'突出端可以与第二核酸分子的平端的互补序列杂交，并且拓扑异构酶可以共价连接ds核酸分子。该方法比先前用于共价连接核酸序列的方法更简单和更有效，并且组合物和试剂盒有助于实施该方法，包括定向连接两个或多个ds核酸分子的方法。

6. 发明申请

US20130004996A1 METHODS AND COMPOSITIONS FOR SYNTHESIS OF NUCLEIC ACID MOLECULES USING MULTIPLERECOGNITION SITES 审中-公开
标题翻译：使用多重分解位点合成核酸分子的方法和组合物
公开(公告)号：US20130004996A1
公开(公告)日：2013-01-03
申请号：US13219440
申请日：2011-08-26
申请人： Jonathan D. CHESNUT , John Carrino , Louis Leong , Knut Madden , Martin A. G. Gleeson , James Fan , Michael A. Brasch , David Cheo , James L. Hartley , Devon R.N. Byrd , Gary F. Temple
发明人： Jonathan D. CHESNUT , John Carrino , Louis Leong , Knut Madden , Martin A. G. Gleeson , James Fan , Michael A. Brasch , David Cheo , James L. Hartley , Devon R.N. Byrd , Gary F. Temple
IPC分类号： C12P19/34 , C12N9/96 , C12N15/66 , C07H21/04
CPC分类号： C12N15/66 , C12N9/90 , C12N15/10 , C12N15/64 , C12N15/902 , C12N2800/108 , C12N2800/30 , C12N2800/70 , C12N2840/20 , C12P19/34 , C12Y599/01
摘要： The present invention provides compositions and methods for recombinational cloning. The compositions include vectors having multiple recombination sites and/or multiple topoisomerase recognition sites. The methods permit the simultaneous cloning of two or more different nucleic acid molecules. In some embodiments the molecules are fused together while in other embodiments the molecules are inserted into distinct sites in a vector. The invention also generally provides for linking or joining through recombination a number of molecules and/or compounds (e.g., chemical compounds, drugs, proteins or peptides, lipids, nucleic acids, carbohydrates, etc.) which may be the same or different. The invention also provides host cells comprising nucleic acid molecules of the invention or prepared according tb the methods of the invention, and also provides kits comprising the compositions, host cells and nucleic acid molecules of the invention, which may be used to synthesize nucleic acid molecules according to the methods of the invention.
摘要翻译：本发明提供了用于重组克隆的组合物和方法。组合物包括具有多个重组位点和/或多个拓扑异构酶识别位点的载体。该方法允许同时克隆两种或更多种不同的核酸分子。在一些实施方案中，分子融合在一起，而在其它实施方案中，分子被插入载体中的不同位点。本发明还通常提供通过重组连接或连接许多可以相同或不同的分子和/或化合物（例如，化合物，药物，蛋白质或肽，脂质，核酸，碳水化合物等）。本发明还提供包含本发明的核酸分子或根据本发明的方法制备的宿主细胞，并且还提供了包含本发明的组合物，宿主细胞和核酸分子的试剂盒，其可用于合成核酸分子根据本发明的方法。

7. 发明申请

US20120129225A1 METHODS AND REAGENTS FOR MOLECULAR CLONING 审中-公开
标题翻译：分子克隆的方法和试剂
公开(公告)号：US20120129225A1
公开(公告)日：2012-05-24
申请号：US13205386
申请日：2011-08-08
申请人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
发明人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
IPC分类号： C12P19/34 , C12N9/96 , C12N15/64
CPC分类号： C12N15/66 , C12N15/10 , C12N15/64
摘要： The present invention provides compositions, methods, and kits for covalently linking nucleic acid molecules. The methods include a strand invasion step, and the compositions and kits are useful for performing such methods. For example, a method of covalently linking double stranded (ds) nucleic acid molecules can include contacting a first ds nucleic acid molecule, which has a topoisomerase linked to a 3′ terminus of one end and has a single stranded 5′ overhang at the same end, with a second ds nucleic acid molecule having a blunt end, such that the 5′ overhang can hybridize to a complementary sequence of the blunt end of the second nucleic acid molecule, and the topoisomerase can covalently link the ds nucleic acid molecules. The methods are simpler and more efficient than previous methods for covalently linking nucleic acid sequences.
摘要翻译：本发明提供用于共价连接核酸分子的组合物，方法和试剂盒。所述方法包括线入侵步骤，组合物和试剂盒对于进行这些方法是有用的。例如，共价连接双链（ds）核酸分子的方法可以包括使第一个ds核酸分子接触，该第一个ds核酸分子具有与一个末端的3'末端连接的拓扑异构酶并且具有相同的单链（5'）突出端末端，具有平末端的第二ds核酸分子，使得5'突出端可以与第二核酸分子的平端的互补序列杂交，并且拓扑异构酶可以共价连接ds核酸分子。该方法比先前用于共价连接核酸序列的方法更简单和更有效。

8. 发明授权

US07528241B2 Methods and reagents for molecular cloning 有权
标题翻译：分子克隆的方法和试剂
公开(公告)号：US07528241B2
公开(公告)日：2009-05-05
申请号：US11053187
申请日：2005-02-07
申请人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
发明人： Jonathan D. Chesnut , Stewart Shuman , Knut R. Madden , John A. Heyman , Robert P. Bennett
IPC分类号： C07H21/02 , C07H21/04 , C12N15/00 , C12Q1/68
CPC分类号： C12N15/66 , C12N15/10 , C12N15/64
摘要： The present invention provides compositions, methods, and kits for covalently linking nucleic acid molecules. The methods include a strand invasion step, and the compositions and kits are useful for performing such methods. For example, a method of covalently linking double stranded (ds) nucleic acid molecules can include contacting a first ds nucleic acid molecule, which has a topoisomerase linked to a 3′ terminus of one end and has a single stranded 5′ overhang at the same end, with a second ds nucleic acid molecule having a blunt end, such that the 5′ overhang can hybridize to a complementary sequence of the blunt end of the second nucleic acid molecule, and the topoisomerase can covalently link the ds nucleic acid molecules. The methods are simpler and more efficient than previous methods for covalently linking nucleic acid sequences, and the compositions and kits facilitate practicing the methods, including methods of directionally linking two or more ds nucleic acid molecules.
摘要翻译：本发明提供用于共价连接核酸分子的组合物，方法和试剂盒。所述方法包括线入侵步骤，组合物和试剂盒对于进行这些方法是有用的。例如，共价连接双链（ds）核酸分子的方法可以包括使第一个ds核酸分子接触，该第一个ds核酸分子具有与一个末端的3'末端连接的拓扑异构酶并且具有相同的单链（5'）突出端末端，具有平末端的第二ds核酸分子，使得5'突出端可以与第二核酸分子的平端的互补序列杂交，并且拓扑异构酶可以共价连接ds核酸分子。该方法比先前用于共价连接核酸序列的方法更简单和更有效，并且组合物和试剂盒有助于实施该方法，包括定向连接两个或多个ds核酸分子的方法。

9. 发明授权

US07033801B2 Compositions and methods for rapidly generating recombinant nucleic acid molecules 有权
标题翻译：用于快速产生重组核酸分子的组合物和方法
公开(公告)号：US07033801B2
公开(公告)日：2006-04-25
申请号：US10014128
申请日：2001-12-07
申请人： John Carrino , James Fan , Robert P. Bennett , Jonathan D. Chesnut , Martin A. Gleeson , Knut R. Madden
发明人： John Carrino , James Fan , Robert P. Bennett , Jonathan D. Chesnut , Martin A. Gleeson , Knut R. Madden
IPC分类号： C12P19/34
CPC分类号： C12P19/34 , C12N9/90 , C12N15/10 , C12N15/64 , C12N15/66 , C12Q1/6855 , C12Q2521/519
摘要： A method of generating a double stranded (ds) recombinant nucleic acid molecule covalently linked in both strands by contacting two or more ds nucleotide sequences with a topoisomerase under conditions such that both termini of at least one end of a first ds nucleotide sequence are covalently linked by the topoisomerase to both termini of at least one end of a second ds nucleotide sequence is provided. Also provided is a method for generating a ds recombinant nucleic acid molecule covalently linked in one strand, by contacting two or more ds nucleotide sequences with a type IA topoisomerase under conditions such that one strand, but not both strands, of one or both ends of a first ds nucleotide sequence are covalently linked by the topoisomerase. Compositions for performing such methods, and compositions generated from such methods also are provided, as are kits containing components useful for conveniently practicing the methods.
摘要翻译：通过使两个或多个ds核苷酸序列与拓扑异构酶接触的条件使得在第一个ds核苷酸序列的至少一个末端的两个末端共价连接的情况下，产生在两条链中共价连接的双链（ds）重组核酸分子的方法提供了拓扑异构酶至第二个ds核苷酸序列的至少一个末端的两个末端。还提供了一种用于产生共价连接在一条链中的ds重组核酸分子的方法，其通过使两个或更多个ds核苷酸序列与IA型拓扑异构酶接触，使得一个或两个末端的一条链但不是两条链第一个ds核苷酸序列由拓扑异构酶共价连接。还提供了用于进行这些方法的组合物，以及由这些方法产生的组合物，以及包含用于方便地实施该方法的组分的试剂盒。

10. 发明授权

US08030066B2 Methods and compositions for synthesis of nucleic acid molecules using multiple recognition sites 有权
标题翻译：使用多个识别位点合成核酸分子的方法和组合物
公开(公告)号：US08030066B2
公开(公告)日：2011-10-04
申请号：US11612445
申请日：2006-12-18
申请人： Jonathan D. Chesnut , John Carrino , Louis Leong , Knut Madden , Martin Gleeson , James Fan , Michael Brasch , David Cheo , James L. Hartley , Devon R. Byrd , Gray F. Temple
发明人： Jonathan D. Chesnut , John Carrino , Louis Leong , Knut Madden , Martin Gleeson , James Fan , Michael Brasch , David Cheo , James L. Hartley , Devon R. Byrd , Gray F. Temple
IPC分类号： C12N15/00
CPC分类号： C12N15/66 , C12N9/90 , C12N15/10 , C12N15/64 , C12N15/902 , C12N2800/108 , C12N2800/30 , C12N2800/70 , C12N2840/20 , C12P19/34 , C12Y599/01
摘要： The present invention provides compositions and methods for recombinational cloning. The compositions include vectors having multiple recombination sites and/or multiple topoisomerase recognition sites. The methods permit the simultaneous cloning of two or more different nucleic acid molecules. In some embodiments the molecules are fused together while in other embodiments the molecules are inserted into distinct sites in a vector. The invention also generally provides for linking or joining through recombination a number of molecules and/or compounds (e.g., chemical compounds, drugs, proteins or peptides, lipids, nucleic acids, carbohydrates, etc.) which may be the same or different. The invention also provides host cells comprising nucleic acid molecules of the invention or prepared according to the methods of the invention, and also provides kits comprising the compositions, host cells and nucleic acid molecules of the invention, which may be used to synthesize nucleic acid molecules according to the methods of the invention.
摘要翻译：本发明提供了用于重组克隆的组合物和方法。组合物包括具有多个重组位点和/或多个拓扑异构酶识别位点的载体。该方法允许同时克隆两种或更多种不同的核酸分子。在一些实施方案中，分子融合在一起，而在其它实施方案中，分子被插入载体中的不同位点。本发明还通常提供通过重组连接或连接许多可以相同或不同的分子和/或化合物（例如，化合物，药物，蛋白质或肽，脂质，核酸，碳水化合物等）。本发明还提供了包含本发明的核酸分子或根据本发明的方法制备的宿主细胞，还提供了包含本发明的组合物，宿主细胞和核酸分子的试剂盒，其可用于合成核酸分子根据本发明的方法。

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