Hangzhou Televector Biomedical Ltd., Co, 310018, Hangzhou, China.
Biotechnol Lett. 2012 Apr;34(4):721-8. doi: 10.1007/s10529-011-0832-0. Epub 2011 Dec 21.
Synthesis of long DNA fragments is often associated with mutations and requires multiple DNA manipulation steps. A novel DNA synthesis method, referred to as patch oligodeoxynucleotide synthesis (POS) to assembly long DNA fragments is presented here. This method involves connection of two types of oligodeoxynucleotides: long constructional oligonucleotides (COs) and short patch oligonucleotides (POs). Long COs were connected by a ligase with the aid of POs, which were complementary to both adjacent COs to help remove secondary structures during assembly. The partial double-stranded DNA template that was formed was then amplified by PCR. Accordingly, we synthesized SV40 polyadenylation signal sequences (187 bp), a codon-optimized yellow fluorescent protein gene (678 bp), and Rattus norvegicus catenin β1 (2,352 bp). This presented method can be broadly applied to synthesize DNA fragments of varying lengths with great convenience.
长 DNA 片段的合成通常与突变有关，需要进行多个 DNA 操作步骤。本文提出了一种新的 DNA 合成方法，称为拼接寡脱氧核苷酸合成（POS）来组装长 DNA 片段。该方法涉及两种类型的寡脱氧核苷酸的连接：长构建性寡核苷酸（COs）和短补丁寡核苷酸（POs）。长 COs 在 POs 的辅助下通过连接酶连接，POs 与两个相邻的 COs 互补，有助于在组装过程中去除二级结构。然后通过 PCR 扩增形成的部分双链 DNA 模板。因此，我们合成了 SV40 多聚腺苷酸化信号序列（187 bp）、密码子优化的黄色荧光蛋白基因（678 bp）和 Rattus norvegicus catenin β1（2352 bp）。该方法可以方便地广泛应用于不同长度 DNA 片段的合成。
