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1.发明授权Methods and compositions for synthesis of nucleic acid molecules using multiple recognition sites 有权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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公开(公告)号：US06905837B2
公开(公告)日：2005-06-14
申请号：US10208557
申请日：2002-07-30
CPC分类号： C12N9/22 , B26D1/0006 , B26D1/08 , B26D2001/006 , B42C5/00 , C12P21/02 , C12Q1/44 , G01N2333/922
摘要： A method is provided for identifying a restriction endonuclease, which includes the steps of (a) screening a target DNA sequence for the presence of known methylase sequence motifs, (b) identifying any open reading frames which lie close to the methylase sequence motifs screened in step (a), and (c) assaying the protein products of these open reading frames for restriction endonuclease activity.Methods for identifying isoschizomers of known restriction endonucleases, which isoschizomers possess a desired physical property, such as thermostability, are also provided by the present invention, as are several novel restriction endonucleases isolated from M. jannaschii, MjaIII and MjaIV. Additionally, a gene was identified that encoded a previously observed endonuclease activity, designated MjaII.Also provided by the present invention are vectors suitable for cloning a DNA sequence encoding a cytotoxic protein, via independent transcription promotors which may be selectively controlled by several conditions. A method for producing these cytotoxic proteins using such vectors is also provided, as are stable clones of PacI and NlaIII.
摘要翻译： 提供了用于鉴定限制性内切核酸酶的方法，其包括以下步骤：（a）筛选靶DNA序列以存在已知的甲基化酶序列基序，（b）鉴定位于接近甲基化酶序列基序的任何开放阅读框， 步骤（a）和（c）测定这些开放阅读框的蛋白质产物用于限制性内切核酸酶活性。 用于鉴定已知限制性内切核酸酶的异构体的方法，其中同分异构体具有期望的物理性质，例如热稳定性，也由本发明提供，以及分离自芒from嗪，MjaIII和MjaIV的几种新的限制性内切核酸酶。 此外，鉴定了编码先前观察到的内切核酸酶活性的基因，命名为MjaII。 本发明还提供了适合于通过独立转录启动子克隆编码细胞毒性蛋白质的DNA序列的载体，其可以通过几种条件选择性地控制。 还提供了使用这种载体产生这些细胞毒性蛋白质的方法，以及PacI和NlaIII的稳定克隆。
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公开(公告)号：US06689573B1
公开(公告)日：2004-02-10
申请号：US09577528
申请日：2000-05-24
IPC分类号： G01N33573
CPC分类号： C12N9/22
摘要： A method is provided for identifying a restriction endonuclease that includes: screening a target DNA sequence for the presence of known methylase sequence motifs, identifying any open reading frames which lie close to the screened methylase sequence motif and assaying the protein products of the open reading frames for restriction endonuclease activity.
摘要翻译： 提供了用于鉴定限制性内切核酸酶的方法，其包括：筛选目标DNA序列以存在已知的甲基化酶序列基序，鉴定位于接近筛选的甲基化酶序列基序的任何开放阅读框，并测定开放阅读框的蛋白质产物 用于限制性内切核酸酶活性。
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公开(公告)号：US06383770B1
公开(公告)日：2002-05-07
申请号：US09486356
申请日：2000-02-25
IPC分类号： C12Q144
CPC分类号： B26D1/0006 , B26D1/08 , B26D2001/006 , B42C5/00 , C12N9/22 , G01N2333/922
摘要： A method is provided for identifying a restriction endonuclease, which includes the steps of (a) screening a target DNA sequence for the presence of known methylase sequence motifs, (b) identifying any open reading frames which lie close to the methylase sequence motifs screened in step (a), and (c) assaying the protein products of these open reading frames for restriction endonuclease activity. Methods for identifying isoschizomers of known restriction endonucleases, which isoschizomers possess a desired physical property, such as thermostability, are also provided by the present invention, as are several novel restriction endonucleases isolated from M. jannaschii, MjaIII and MjaIV. Additionally, a gene was identified that encoded a previously observed endonuclease activity, designated MjaII. Also provided by the present invention are vectors suitable for cloning a DNA sequence encoding a cytotoxic protein, via independent transcription promotors which may be selectively controlled by several conditions. A method for producing these cytotoxic proteins using such vectors is also provided, as are stable clones of PacI and NlaIII.
摘要翻译： 提供了用于鉴定限制性内切核酸酶的方法，其包括以下步骤：（a）筛选靶DNA序列以存在已知的甲基化酶序列基序，（b）鉴定位于接近甲基化酶序列基序的任何开放阅读框， 步骤（a）和（c）测定这些开放阅读框的蛋白质产物用于限制性内切核酸酶活性。 用于鉴定已知限制性内切核酸酶的异构体的方法，其中同分异构体具有期望的物理性质，例如热稳定性，也由本发明提供，以及从分枝杆菌，MjaIII和MjaIV分离的几种新的限制性内切核酸酶。 此外，鉴定了编码先前观察到的内切核酸酶活性的基因，命名为MjaII。 本发明还提供了适合于通过独立转录启动子克隆编码细胞毒性蛋白质的DNA序列的载体，其可以通过几种条件选择性地控制。 还提供了使用这种载体产生这些细胞毒性蛋白质的方法，以及PacI和NlaIII的稳定克隆。
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