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1. 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。

2. 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。

3. 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。

4. 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。

5. 发明授权

US07078501B2 Topoisomerase linker-mediated amplification methods 失效
标题翻译：拓扑异构酶接头介导的扩增方法
公开(公告)号：US07078501B2
公开(公告)日：2006-07-18
申请号：US09792875
申请日：2001-02-23
申请人： John A. Heyman , Gabor Szalai , Michael Felder , Knut R. Madden
发明人： John A. Heyman , Gabor Szalai , Michael Felder , Knut R. Madden
IPC分类号： C07H21/04 , C12Q1/68 , C12P19/34
CPC分类号： C12Q1/6869 , C12N15/10 , C12Q1/6855 , C12Q2521/519 , C12Q2525/191 , C12Q2531/113
摘要： A Vaccinia topoisomerase-adapted linker, which contains an oligonucleotide primer binding site of known sequence, useful for specifically joining the linker to the end of a polynucleotide of unknown sequence and allowing subsequent PCR amplification of the polynucleotide or DNA is provided. Kits containing the invention Vaccinia topoisomerase-adapted linker and one or more linker-specific oligonucleotides for annealing to the linker in PCR are also provided. In addition, the invention provides methods for using the invention linkers in linker-mediated PCR amplification procedures for isolation and optional sequencing of isolated PCR amplification products.
摘要翻译：提供了一种具有已知序列的寡核苷酸引物结合位点的牛痘拓扑异构酶适配接头，其可用于将接头特异性连接到未知序列的多核苷酸的末端并允许随后的多核苷酸或DNA的PCR扩增。还提供了含有本发明的Vaccinia拓扑异构酶修饰接头和用于在PCR中对接头退火的一个或多个接头特异性寡核苷酸的试剂盒。此外，本发明提供了使用本发明接头在接头介导的PCR扩增程序中用于分离和任选测序分离的PCR扩增产物的方法。

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