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    • 6. 发明申请
    • Compositions and Methods for High Fidelity Assembly of Nucleic Acids
    • 核酸高保真组装的组成和方法
    • US20150203839A1
    • 2015-07-23
    • US14241242
    • 2012-08-23
    • Joseph JacobsonDaniel SchindlerScott S. Lawton
    • Joseph JacobsonDaniel SchindlerScott S. Lawton
    • C12N15/10
    • C12N15/1031C12N15/10C12N15/1027C12N15/1089C12N15/1093C12N15/66C12P19/34C12Q2521/301C12Q2521/501
    • Aspects of the invention relate to methods, compositions and algorithms for designing and producing a target nucleic acid. The method can include: (1) providing a plurality of blunt-end double-stranded nucleic acid fragments having a restriction enzyme recognition sequence at both ends thereof; (2) producing via enzymatic digestion a plurality of cohesive-end double-stranded nucleic acid fragments each having two different and non-complementary overhangs; (3) ligating the plurality of cohesive-end double-stranded nucleic acid fragments with a ligase; and (4) forming a linear arrangement of the plurality of cohesive-end double-stranded nucleic acid fragments, wherein the unique arrangement comprises the target nucleic acid. In certain embodiments, the plurality of blunt-end double-stranded nucleic acid fragments can be provided by: releasing a plurality of oligonucleotides synthesized on a solid support; and synthesizing complementary strands of the plurality of oligonucleotides using a polymerase based reaction.
    • 本发明的方面涉及用于设计和产生靶核酸的方法,组合物和算法。 该方法可以包括:(1)提供多个在其两端具有限制酶识别序列的平末端双链核酸片段; (2)通过酶消化产生多个具有两个不同和非互补突出端的粘性末端双链核酸片段; (3)用连接酶连接多个内聚端双链核酸片段; 和(4)形成多个内聚端双链核酸片段的线性排列,其中独特的排列包含靶核酸。 在某些实施方案中,多个平末端双链核酸片段可以通过以下方式提供:释放在固体支持物上合成的多种寡核苷酸; 并使用基于聚合酶的反应合成多个寡核苷酸的互补链。