Rudenko G N, Rommens C M, Nijkamp H J, Hille J
Free University, Department of Genetics, Amsterdam, Netherlands.
Plant Mol Biol. 1993 Feb;21(4):723-8. doi: 10.1007/BF00014557.
We describe a novel modification of the polymerase chain reaction for efficient in vitro amplification of genomic DNA sequences flanking short stretches of known sequence. The technique utilizes a target enrichment step, based on the selective isolation of biotinylated fragments from the bulk of genomic DNA on streptavidin-containing support. Subsequently, following ligation with a second universal linker primer, the selected fragments can be amplified to amounts suitable for further molecular studies. The procedure has been applied to recover T-DNA flanking sequences in transgenic tomato plants which could subsequently be used to assign the positions of T-DNA to the molecular map of tomato. The method called supported PCR (sPCR) is a simple and efficient alternative to techniques used in the isolation of specific sequences flanking a known DNA segment.
我们描述了一种聚合酶链反应的新改良方法，用于高效体外扩增已知序列短片段侧翼的基因组DNA序列。该技术利用了一个靶标富集步骤，此步骤基于从含链霉亲和素的载体上的大量基因组DNA中选择性分离生物素化片段。随后，在用第二个通用接头引物连接后，所选片段可被扩增至适合进一步分子研究的量。该程序已应用于回收转基因番茄植株中的T-DNA侧翼序列，这些序列随后可用于将T-DNA的位置定位到番茄的分子图谱上。这种称为支持PCR（sPCR）的方法是分离已知DNA片段侧翼特定序列所使用技术的一种简单而有效的替代方法。
