Herrmann R, Neugebauer K, Pirkl E, Zentgraf H, Schaller H
Mol Gen Genet. 1980 Jan;177(2):231-42. doi: 10.1007/BF00267434.
Single-stranded DNA vectors were constructed in vitro by insertion of various DNA fragments into the Intergenic Region of the single-stranded DNA phage fd. These inserts introduce into the phage genome unique cleavage sites for restriction nucleases which are suited for sticky joining in cloning experiments. Since these sites are usually located within genes coding for antibiotic resistance, inactivation of a resistance gene by insertion can be used as a marker for the successful cloning of a DNA fragment. Resistance genes also allow to select for recombinant DNA phages and to minimize the loss of DNA inserts which otherwise becomes significant above an insert size of about one kb. Cloning of several DNA fragments is described and strand separation of double-stranded DNA fragments by means of cloning into fd DNA is given as an example for application of single-stranded DNA vectors.
通过将各种DNA片段插入单链DNA噬菌体fd的基因间隔区，在体外构建了单链DNA载体。这些插入片段在噬菌体基因组中引入了限制核酸酶的独特切割位点，这些位点适用于克隆实验中的粘性连接。由于这些位点通常位于编码抗生素抗性的基因内，通过插入使抗性基因失活可作为DNA片段成功克隆的标记。抗性基因还可用于选择重组DNA噬菌体，并最大限度地减少DNA插入片段的丢失，否则在插入片段大小约为1 kb以上时，这种丢失会变得很显著。本文描述了几个DNA片段的克隆，并以通过克隆到fd DNA中实现双链DNA片段的链分离为例，介绍了单链DNA载体的应用。
