Combinatorial approaches in directed evolution were proven to be more efficient for exploring sequence space and innovating function of protein. Here, we presented the modular assembly of secondary structures (MASS) for constructing a combinatorial library. In this approach, secondary structure elements were extracted from natural existing protein. The common linkers were flanking secondary structure elements, and then secondary structure elements were digested by Hinf I restriction endonuclease that was used in the construction of combinatorial library for the first time. The digested DNA fragments were randomly ligated in the sense orientation, then in sequence to be amplified by PCR and transformation. This approach showed that different DNA fragments without homologous sequences could be randomly assembled to create significant sequence space. With the structure analysis of recombinants, it would be beneficial to the rational design, even to the design of protein de novo, and to evolve any genetic part or circuit.

中文翻译：

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基于限制酶介导的模块化装配的组合库。

事实证明，定向进化中的组合方法对于探索序列空间和创新蛋白质功能更为有效。在这里，我们介绍了用于构造组合库的二级结构的模块化组装（MASS）。在这种方法中，二级结构元素是从天然存在的蛋白质中提取的。常见的接头位于二级结构元件的侧翼，然后通过Hinf I限制性核酸内切酶消化二级结构元件，该酶首次用于组合文库的构建。消化后的DNA片段以有义方向随机连接，然后按顺序通过PCR和转化进行扩增。该方法表明，可以将无同源序列的不同DNA片段随机组装起来，以创建明显的序列空间。