Biotechnology Department, The School of Medical and Life Science, University of Jinan, Jinan, P. R. China.
Electrophoresis. 2010 Sep;31(17):2929-35. doi: 10.1002/elps.201000222.
With a novel and universal strategy for the cloning of multiple DNA fragments, a complex synthetic vector (pVEC100), harboring the target DNA fragments in conventional 100-bp DNA ladder, was constructed for efficient and large-scale production of 100-bp DNA marker through bacteria fermentation, plasmid extraction and restrictive digestion. Since the restrictive digestion of complex vectors yields insufficient small DNA fragments, an innovative PCR model was developed as an alternative. The PCR model comprised a specially designed template vector and a unit amplification model for producing groups of small DNA fragments. The unit amplification model improved the efficiency of the PCR protocol and made it more economical and easier for small DNA fragment amplification. The approach presented in this paper--a unit cloning model for constructing complex synthetic vectors combined with the modular design of unit amplification by PCR--is a powerful method for preparing small DNA fragments of DNA molecular weight standards.
采用一种新颖而通用的方法对多个 DNA 片段进行克隆，构建了一个复杂的合成载体（pVEC100），该载体以常规的 100-bp DNA 梯状排列目标 DNA 片段，通过细菌发酵、质粒提取和限制性消化，高效、大规模地生产 100-bp DNA 标记物。由于复杂载体的限制性消化会产生不足够的小 DNA 片段，因此开发了一种创新的 PCR 模型作为替代方法。该 PCR 模型包括一个专门设计的模板载体和一个用于产生多组小 DNA 片段的单元扩增模型。单元扩增模型提高了 PCR 方案的效率，使其更经济实惠，更便于小 DNA 片段的扩增。本文提出的方法——一种用于构建复杂合成载体的单元克隆模型，结合通过 PCR 进行单元扩增的模块化设计——是一种用于制备 DNA 分子量标准的小 DNA 片段的强大方法。
