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    • 4. 发明申请
    • Improved Methods for Rapid Gene Synthesis
    • 改进的快速基因合成方法
    • US20120156731A1
    • 2012-06-21
    • US12052156
    • 2008-03-20
    • Yanyi HuangJames BergerStephen Quake
    • Yanyi HuangJames BergerStephen Quake
    • C12P19/34C07H21/04C12M1/00C12N9/00
    • C12Y605/01001C12P19/34C12Y605/01002
    • Disclosed are methods and materials for assembling long polynucleotides from synthetic oligonucleotides. The use of synthetic oligonucleotides permits non-natural design of sequences. The oligonucleotides used for construction may be relatively short, according to practicalities of nucleotide synthesis. They are assembled using a ligase which is operative over a range of temperatures, i.e., is thermostable. The method and oligonucleotides are designed such that the melting temperature of the strands to be hybridized is set at a number of selected specific temperatures for each group of oligonucleotides to be hybridized and ligated. Hybridization and ligation take place at or near the melting temperature, so that each succeeding ligation is governed by a temperature that will prevent hybridization if any mismatches are present.
    • 公开了用于从合成寡核苷酸组装长多核苷酸的方法和材料。 合成寡核苷酸的使用允许序列的非自然设计。 根据核苷酸合成的实用性,用于构建的寡核苷酸可能相对较短。 它们使用在一定温度范围内操作的连接酶组装,即是耐热的。 设计该方法和寡核苷酸使待杂交的链的解链温度被设定为要混合并连接的每组寡核苷酸的选定的特定温度的数目。 杂交和连接在熔融温度或接近熔化温度下进行,因此每次后续连接都由温度控制,如果存在任何错配,将会阻止杂交。